How the immune mousetrap works: Structural evidence for the immunomodulatory action of a peptide from influenza NS1 protein.

One of the critical stages of the T-cell immune response is the dimerization of the intramembrane domains of T-cell receptors (TCR). Structural similarities between the immunosuppressive domains of viral proteins and the transmembrane domains of TCR have led several authors to hypothesize the mechanism of immune response suppression by highly pathogenic viruses: viral proteins embed themselves in the membrane and act on the intramembrane domain of the TCRalpha subunit, hindering its functional oligomerization. It has also been suggested that this mechanism is used by influenza A virus in NS1-mediated immunosuppression. We have shown that the peptide corresponding to the primary structure of the potential immunosuppressive domain of NS1 protein (G51) can reduce concanavalin A-induced proliferation of PBMC cells, as well as in vitro, G51 can affect the oligomerization of the core peptide corresponding to the intramembrane domain of TCR, using AFM and small-angle neutron scattering. The results obtained using in cellulo and in vitro model systems suggest the presence of functional interaction between the NS1 fragment and the intramembrane domain of the TCR alpha subunit. We have proposed a possible scheme for such interaction obtained by computer modeling. This suggests the existence of another NS1-mediated mechanism of immunosuppression in influenza.

Variability of internal standard method calibration factors estimated with a multifactorial Taguchi experiment in the analysis of alcoholic products by gas chromatography with flame ionization detection.

This study compares the variability of relative response factors (RRFs) using Taguchi's multifactorial analysis for two internal standard (IS) methods in gas chromatography (GC) for quality control of alcoholic products. Methods where either ethanol or pentan-1-ol was used as an IS were compared. For ten volatile substances prescribed by legislation, the RRFs of both methods were compared under 27 different experimental conditions. The influence of parameters (control factors) such as ethanol content in the matrix, analyte concentration, injected volume, injector temperature, split ratio, and flame ionization detector temperature was evaluated. The selected control factors had values at one of the three levels to cover the commonly used ranges of their settings in the measuring system and to characterize the majority of alcoholic products commonly analyzed in practice. The obtained results showed that the biggest differences in the variability of the results between the two methods were found for the most strictly controlled substances in alcoholic products, acetaldehyde, and methanol, where the application of ethanol as an IS provides clearly better results. For both methods, the way control factors affect the repeatability of GC measurements expressed in the form of relative deviation was also evaluated.

Interaction of Lactoferrin with Unsaturated Fatty Acids: In Vitro and In Vivo Study of Human Lactoferrin/Oleic Acid Complex Cytotoxicity.

As shown recently, oleic acid (OA) in complex with lactoferrin (LF) causes the death of cancer cells, but no mechanism(s) of that toxicity have been disclosed. In this study, constitutive parameters of the antitumor effect of LF/OA complex were explored. Complex LF/OA was prepared by titrating recombinant human LF with OA. Spectral analysis was used to assess possible structural changes of LF within its complex with OA. Structural features of apo-LF did not change within the complex LF:OA = 1:8, which was toxic for hepatoma 22a cells. Cytotoxicity of the complex LF:OA = 1:8 was tested in cultured hepatoma 22a cells and in fresh erythrocytes. Its anticancer activity was tested in mice carrying hepatoma 22a. In mice injected daily with LF-8OA, the same tumor grew significantly slower. In 20% of animals, the tumors completely resolved. LF alone was less efficient, i.e., the tumor growth index was 0.14 for LF-8OA and 0.63 for LF as compared with 1.0 in the control animals. The results of testing from 48 days after the tumor inoculation showed that the survival rate among LF-8OA-treated animals was 70%, contrary to 0% rate in the control group and among the LF-treated mice. Our data allow us to regard the complex of LF and OA as a promising tool for cancer treatment.

Predictive Value of Biomechanical Mapping for Pelvic Organ Prolapse Surgery.

OBJECTIVE: This study examined biomechanical changes in pelvic floor after urogynecological surgery. METHODS: This multisite clinical study was designed to explore changes in tissue elasticity, pelvic support, and certain functions (contractive strength, muscle relaxation speed, muscle motility) after pelvic organ prolapse (POP) surgery. A biomechanical mapping of the pelvic floor was performed before and 4 to 6 months after the surgery. The biomechanical data for 52 parameters were acquired by vaginal tactile imaging for manually applied deflection pressures to vaginal walls and pelvic muscle contractions. The two-sample t-test (P < 0.05) was used to test the null hypothesis that presurgery data in group 1 (positive parameter change after surgery) and presurgery data in group 2 (negative parameter change after surgery) belonged to the same distribution. RESULTS: A total of 78 subjects with 255 surgical procedures were analyzed across 5 participating clinical sites. All 52 t-tests for group 1 versus group 2 had P value in the range from 4.0 × 10-10 to 4.3 × 10-2 associating all of the 52 parameter changes after surgery with the presurgical conditions. The P value of before and after surgery correlation ranged from 3.7 × 10-18 to 1.6 × 10-2 for 50 of 52 tests, with Pearson correlation coefficient ranging from -0.79 to -0.27. Thus, vaginal tactile imaging parameters strongly correlated weak pelvic floor presurgery with the positive POP surgery outcome of improved biomechanical properties. CONCLUSIONS: Pelvic organ prolapse surgery, in general, improves the biomechanical conditions and integrity of the weak pelvic floor. The proposed biomechanical parameters can predict changes resulting from POP surgery.

Effect of alpha-lactalbumin and lactoferrin oleic acid complexes on chromatin structural organization.

This work focuses on the study of multimeric alpha-lactalbumin oleic acid and lactoferrin oleic acid complexes. The purpose of the research is to study possible mechanisms involved in their pro-apoptotic activities, as seen in some tumor cell cultures. Complexes featuring oleic acid (OA) with human alpha-lactalbumin (hAl) or with bovine alpha-lactalbumin (bAl), and human lactoferrin (hLf) were investigated using small-angle neutron scattering (SANS). It was shown that while alpha-lactalbumin protein complexes were formed on the surface of polydisperse OA micelles, the lactoferrin complexes comprised a monodisperse system of nanoscale particles. Both hAl and hLf complexes appeared to interact with the chromatin of isolated nuclei affecting chromatin structural organization. The possible roles of these processes in the specific anti-tumor activity of these complexes are discussed.

Changing times: Fluorescence-lifetime analysis of amyloidogenic SF-IAPP fusion protein.

In a number of conformational diseases, intracellular accumulation of proteins bearing non-native conformations occurs. The search for compounds that are capable of hindering the formation and accumulation of toxic protein aggregates and fibrils is an urgent task. Present fluorescent methods of fibrils' detection prevent simple real-time observations. We suppose to use green fluorescent protein fused with target protein and fluorescence lifetime measurement technique for this purpose. The recombinant proteins analyzed were produced in E. coli. Mass spectrometry was used for the primary structure of the recombinant proteins and post-translational modifications identification. The fluorescence lifetime of the superfolder green fluorescent protein (SF) and the SF protein fused with islet amyloid polypeptide (SF-IAPP) were studied in polyacrylamide gel using Fluorescent-Lifetime Imaging Microscopy (FLIM). It was shown that the SF average fluorescence lifetime in gel slightly differs from that of the SF-IAPP monomer under these conditions. SF-IAPP does not lose the ability to form amyloid-like fibrils. Under the same conditions (in polyacrylamide gel), SF and SF-IAPP monomers have similar fluorescence time characteristics and the average fluorescence lifetime of SF-IAPP in fibrils significantly decreases. We propose the application of FLIM to the measurement of average fluorescence lifetimes of fusion proteins (amyloidogenic protein-SF) in the context of studies using cellular models of conformational diseases.

Triazavirine supramolecular complexes as modifiers of the peptide oligomeric structure.

In this study, we present molecular dynamics simulations of the antiviral drug triazavirine, that affects formation of amyloid-like fibrils of the model peptide (SI). According to our simulations, triazavirine is able to form linear supramolecular structures which can act as shields and prevent interactions between SI monomers. This model, as validated by simulations, provides an adequate explanation of triazavirine's mechanism of action as it pertains to SI peptide fibril formation.

Quantitative Assessment and Interpretation of Vaginal Conditions.

INTRODUCTION: Few means exist to provide quantitative and reproducible assessment of vaginal conditions from biomechanical and functional standpoints. AIM: To develop a new approach for quantitative biomechanical characterization of the vagina. METHODS: Vaginal tactile imaging (VTI) allows biomechanical assessment of soft tissue and function along the entire length of the anterior, posterior, and lateral vaginal walls. This can be done at rest, with applied vaginal deformation, and with pelvic muscle contraction. RESULTS: Data were analyzed for 42 subjects with normal pelvic floor support from an observational case-controlled clinical study. The average age was 52 years (range = 26-90 years). We introduced 8 VTI parameters to characterize vaginal conditions: (i) maximum resistance force to insertion (newtons), (ii) insertion work (millijoules), (iii) maximum stress-to-strain ratio (elasticity; kilopascals per millimeter), (iv) maximum pressure at rest (kilopascals), (v) anterior-posterior force at rest (newtons), (vi) left-right force at rest (newtons), (vii) maximum pressure at muscle contraction (kilopascals), and (viii) muscle contraction force (newtons). We observed low to moderate correlation of these parameters with subject age and no correlation with subject weight. 6 of 8 parameters demonstrated a P value less than .05 for 2 subject subsamples divided by age (≤52 vs >52 years), which means 6 VTI parameters change with age. CONCLUSIONS: VTI allows biomechanical and functional characterization of the vaginal conditions that can be used for (i) understanding "normal" vaginal conditions, (ii) quantification of the deviation from normality, (iii) personalized treatment (radiofrequency, laser, or plastic surgery), and (iv) assessment of the applied treatment outcome. Egorov V, Murphy M, Lucente V, et al. Quantitative Assessment and Interpretation of Vaginal Conditions. Sex Med 2018;6:39-48.

Objective Differences in Colonoscopy Technique Between Trainee and Expert Endoscopists Using the Colonoscopy Force Monitor.

BACKGROUND: Learning to perform colonoscopy safely and effectively is central to gastroenterology fellowship programs. The application of force to the colonoscope is an important part of colonoscopy technique. AIMS: We compared force application during colonoscopy between novice and expert endoscopists using a novel device to determine differences in colonoscopy technique. METHODS: This is an observational cohort study designed to compare force application during colonoscopy between novice and experienced trainees, made up of gastroenterology fellows from two training programs, and expert endoscopists from both academic and private practice settings. RESULTS: Force recordings were obtained for 257 colonoscopies by 37 endoscopists, 21 of whom were trainees. Experts used higher average forward forces during insertion compared to all trainees and significantly less clockwise torque compared to novice trainees. CONCLUSIONS: We present significant, objective differences in colonoscopy technique between novice trainees, experienced trainees, and expert endoscopists. These findings suggest that the colonoscopy force monitor is an objective tool for measuring proficiency in colonoscopy. Furthermore, the device may be used as a teaching tool in training and continued medical education programs.

Peptide-Induced Amyloid-Like Conformational Transitions in Proteins.

Changes in protein conformation can occur both as part of normal protein functioning and during disease pathogenesis. The most common conformational diseases are amyloidoses. Sometimes the development of a number of diseases which are not traditionally related to amyloidoses is associated with amyloid-like conformational transitions of proteins. Also, amyloid-like aggregates take part in normal physiological processes such as memorization and cell signaling. Several primary structural features of a protein are involved in conformational transitions. Also the protein proteolytic fragments can cause the conformational transitions in the protein. Short peptides which could be produced during the protein life cycle or which are encoded by short open reading frames can affect the protein conformation and function.

Awake video-assisted thoracic surgery in acute infectious pulmonary destruction.

BACKGROUND: Many of thoracic minimally invasive interventions have been proven to be possible without general anesthesia. This article presents results of video-assisted thoracic surgery (VATS) application under local anesthesia in patients with lung abscesses and discusses its indications in detail. METHODS: The study involved prospective analysis of treatment outcomes for all acute infectious pulmonary destruction (AIPD) patients undergoing VATS under local anesthesia and sedation since January 1, 2010, till December 31, 2013. Patients with pulmonary destruction cavity at periphery of large size (>5 cm) underwent non-intubated video abscessoscopy (NIVAS). Patients with pyopneumothorax (lung abscess penetration into pleural cavity) underwent non-intubated video thoracoscopy (NIVTS). Indications for NIVAS and NIVTS were as follows: cavity debridement and washing, necrotic sequestra removal, adhesion split, biopsy. All interventions were done under local anesthesia and sedation without trachea intubation and epidural anesthesia. RESULTS: Sixty-five enrolled patients had 42 NIVAS and 32 NIVTS interventions, nine patients underwent two surgeries. None of the patients required trachea intubation or epidural anesthesia. In none of our cases with conversion to thoracotomy was required. Post-surgical complications developed after 11 interventions (13%): subcutaneous emphysema (five cases), chest wall phlegmon (three cases), pulmonary bleeding (two cases), and pneumothorax (one case). One patient died due to the main disease progression. In 50 patients NIVAS and NIVTS were done within 5 to 8 days after abscess/pleural cavity draining, while in other 15 patients-immediately prior to draining; both pulmonary bleeding episodes and all cases of chest wall phlegmon took place in the latter group. CONCLUSIONS: NIVAS and NIVTS under local anesthesia and sedation are well tolerated by patients, safe and should be used more often in AIPD cases. Timing of NIVAS and NIVTS procedures was found to be of paramount importance for ensuring complete therapeutic effectiveness.

Effect of propofol anesthesia on force application during colonoscopy.

BACKGROUND: Sedation is frequently used during colonoscopy to control patient discomfort and pain. Propofol is associated with a deeper level of sedation than is a combination of a narcotic and sedative hypnotic and, therefore, may be associated with an increase in force applied to the colonoscope to advance and withdraw the instrument. OBJECTIVE: To compare force application to the colonoscope insertion tube during propofol anesthesia and moderate sedation. DESIGN: An observational cohort study of 13 expert and 12 trainee endoscopists performing colonoscopy in 114 patients. Forces were measured by using the colonoscopy force monitor, which is a wireless, handheld device that attaches to the insertion tube of the colonoscope. SETTING: Community ambulatory surgery center and academic gastroenterology training programs. PATIENTS: Patients undergoing routine screening or diagnostic colonoscopy with complete segment force recordings. MAIN OUTCOME MEASUREMENTS: Axial and radial forces and examination time. RESULTS: Axial and radial forces increase and examination time decreases significantly when propofol is used as the method of anesthesia. LIMITATIONS: Small study, observational design, nonrandomized distribution of sedation type and experience level, different instrument type and effect of prototype device on insertion tube manipulation. CONCLUSIONS: Propofol sedation is associated with a decrease in examination time and an increase in axial and radial forces used to advance the colonoscope.

Vaginal tactile imaging.

Changes in the elasticity of the vaginal walls, connective support tissues, and muscles are thought to be significant factors in the development of pelvic organ prolapse, a highly prevalent condition affecting at least 50% of women in the United States during their lifetimes. It creates two predominant concerns specific to the biomechanical properties of pelvic support tissues: how does tissue elasticity affect the development of pelvic organ prolapse and how can functional elasticity be maintained through reconstructive surgery. We designed a prototype of vaginal tactile imager (VTI) for visualization and assessment of elastic properties of pelvic floor tissues. In this paper, we analyze applicability of tactile imaging for evaluation of reconstructive surgery results and characterization of normal and pelvic organ prolapse conditions. A pilot clinical study with 13 patients demonstrated that VTI allows imaging of vaginal walls with increased rigidity due to implanted mesh grafts following reconstructive pelvic surgery and VTI has the potential for prolapse characterization and detection.

Characterization of forces applied by endoscopists during colonoscopy by using a wireless colonoscopy force monitor.

BACKGROUND: To perform a colonoscopy, the endoscopist maneuvers the colonoscope through a series of loops by applying force to the insertion tube. Colonoscopy insertion techniques are operator dependent but have never been comprehensively quantified. OBJECTIVE: To determine whether the Colonoscopy Force Monitor (CFM), a device that continually measures force applied to the insertion tube, can identify different force application patterns among experienced endoscopists. DESIGN: Observational study of 6 experienced endoscopists performing routine diagnostic and therapeutic colonoscopy in 30 patients. SETTING: Outpatient ambulatory endoscopy center. PATIENTS: Adult male and female patients between 30 and 75 years of age undergoing routine colonoscopy. INTERVENTIONS: CFM monitoring of force applied to the colonoscope insertion tube during colonoscopy. MAIN OUTCOME MEASUREMENTS: Maximum and mean linear and torque force, time derivative of force, combined linear and torque vector force, and total manipulation time. RESULTS: The CFM demonstrates differences among endoscopists for maximum and average push/pull and mean torque forces, time derivatives of force, combined push/torque force vector, and total manipulation time. Endoscopists could be grouped by force application patterns. LIMITATIONS: Only experienced endoscopists using conscious sedation in the patients were studied. Sample size was 30 patients. CONCLUSIONS: This study demonstrates that CFM allows continuous force monitoring, characterization, and display of similarities and differences in endoscopic technique. CFM has the potential to facilitate training by enabling trainees to assess, compare, and quantify their techniques and progress.

Mechanical imaging in medical applications.

Mechanical Imaging (MI), a.k.a. tactile imaging or stress imaging, is a branch of Elasticity Imaging, a medical diagnostic technique based on the visualization of tissue internal structures in terms of their elasticity modulus. During the last decade, numerous methods and devices have been developed implementing MI technology in various medical applications, such as the visualization and evaluation of prostate conditions, breast cancer screening, the differentiation of benign and malignant lesions, and the characterization of vaginal wall elasticity. This paper presents an overview of MI technology and its applications, strengths and limitations. Results of laboratory and clinical studies clearly indicate that Mechanical Imaging devices have the potential to be used as a cost effective means for cancer screening as well as diagnostics of various diseases accompanied by changes of mechanical properties of soft tissues.

Differentiation of benign and malignant breast lesions by mechanical imaging.

Mechanical imaging yields tissue elasticity map and provides quantitative characterization of a detected pathology. The changes in the surface stress patterns as a function of applied load provide information about the elastic composition and geometry of the underlying tissue structures. The objective of this study is the clinical evaluation of breast mechanical imager for breast lesion characterization and differentiation between benign and malignant lesions. The breast mechanical imager includes a probe with pressure sensor array, an electronic unit providing data acquisition from the pressure sensors and communication with a touch-screen laptop computer. We have developed an examination procedure and algorithms to provide assessment of breast lesion features such as hardness related parameters, mobility, and shape. A statistical Bayesian classifier was constructed to distinguish between benign and malignant lesions by utilizing all the listed features as the input. Clinical results for 179 cases, collected at four different clinical sites, have demonstrated that the breast mechanical imager provides a reliable image formation of breast tissue abnormalities and calculation of lesion features. Malignant breast lesions (histologically confirmed) demonstrated increased hardness and strain hardening as well as decreased mobility and longer boundary length in comparison with benign lesions. Statistical analysis of differentiation capability for 147 benign and 32 malignant lesions revealed an average sensitivity of 91.4% and specificity of 86.8% with a standard deviation of +/-6.1%. The area under the receiver operating characteristic curve characterizing benign and malignant lesion discrimination is 86.1% with the confidence interval ranging from 80.3 to 90.9%, with a significance level of P = 0.0001 (area = 50%). The multisite clinical study demonstrated the capability of mechanical imaging for characterization and differentiation of benign and malignant breast lesions. We hypothesize that the breast mechanical imager has the potential to be used as a cost effective device for cancer diagnostics that could reduce the benign biopsy rate, serve as an adjunct to mammography and to be utilized as a screening device for breast cancer detection.

Prostate mechanical imaging: a new method for prostate assessment.

OBJECTIVES: To evaluate the ability of prostate mechanical imaging (PMI) technology to provide an objective and reproducible image and to assess the prostate nodularity. METHODS: We evaluated the PMI device developed by Artann Laboratories in a pilot clinical study. For the 168 patients (ages 44 to 94) who presented to an urologist for prostate evaluation, PMI-produced images and assessment of prostate size, shape, consistency/hardness, mobility, and nodularity were compared with digital rectal examination (DRE) findings. The PMI and DRE results were further tested for correlation against a transrectal ultrasound of the prostate (TRUS) guided biopsy for a subgroup of 21 patients with an elevated prostate-specific antigen level. RESULTS: In 84% of the cases, the PMI device was able to reconstruct three-dimensional (3D) and 2D cross-sectional images of the prostate. The PMI System and DRE pretests were able to determine malignant nodules in 10 and 6 patients, respectively, of the 13 patients with biopsy-confirmed malignant inclusions. The PMI System findings were consistent with all 8 biopsy negative cases, whereas the DRE had 1 abnormal reading for this group. The correlation between PMI and DRE detection of palpable nodularity was 81%, as indicated by the area under the receiver operating characteristic curve. Estimates of the prostate size provided by PMI and DRE were statistically significantly correlated. CONCLUSIONS: The PMI has the potential to enable a physician to obtain, examine, and store a 3D image of the prostate based on mechanical and geometrical characteristics of the gland and its internal structures.

Prostate mechanical imaging: 3-D image composition and feature calculations.

We have developed a method and a device entitled prostate mechanical imager (PMI) for the real-time imaging of prostate using a transrectal probe equipped with a pressure sensor array and position tracking sensor. PMI operation is based on measurement of the stress pattern on the rectal wall when the probe is pressed against the prostate. Temporal and spatial changes in the stress pattern provide information on the elastic structure of the gland and allow two-dimensional (2-D) and three-dimensional (3-D) reconstruction of prostate anatomy and assessment of prostate mechanical properties. The data acquired allow the calculation of prostate features such as size, shape, nodularity, consistency/hardness, and mobility. The PMI prototype has been validated in laboratory experiments on prostate phantoms and in a clinical study. The results obtained on model systems and in vivo images from patients prove that PMI has potential to become a diagnostic tool that could largely supplant DRE through its higher sensitivity, quantitative record storage, ease-of-use and inherent low cost.

Thoracoscopic collagen pleurodesis in the treatment of malignant pleural effusions.

OBJECTIVE: Pleurodesis is of a potential benefit in pleural carcinomatosis and symptomatic malignant effusions, but the best way of achieving this is still uncertain. The aim of this prospective study was to analyse the results of pleurodesis after intra-pleural thoracoscopic administration of collagen powder. METHODS: 45 patients (19 men and 26 women; median age of 64 years, range from 36 to 73 years) with malignant pleural effusions underwent thoracoscopic collagen pleurodesis. The procedure involved thoracoscopic drainage of pleural effusion and intra-pleural insufflation of 1 g of bovine dermal collagen powder under general anaesthesia. Assessment of the immediate side effects and pH estimation of drained pleural fluid took place whilst inpatient. The patients were subsequently followed up for 1 year at 3-monthly intervals including outpatient clinical review and chest radiography. Prognostic value of pleural fluid pH in relation to the outcome of pleurodesis and patients' survival was statistically analysed. RESULTS: The procedure was well tolerated and there were no serious complications or deaths. Thoracoscopic collagen pleurodesis resulted in immediate resolution of malignant pleural effusion and all patients remained free of re-accumulated fluid for at least 1 month. Only 5 (11%) patients later developed recurrent effusion and required its repeat drainage at some point during the follow-up period. In the vast majority (89%) patients, thoracoscopic collagen pleurodesis proved successful in complete and permanent resolution of pleural fluid collection. Acid medium (pH < 7.3) of plural fluid was associated with poor survival (P < 0.05), but did not influence the clinical and radiological outcome of collagen pleurodesis (P > 0.05). CONCLUSIONS: Thoracoscopic collagen pleurodesis is a simple and effective method of treatment of malignant pleural effusions.