Evaluation of Antitumoral activity of bone targeted drugs/conventional chemotherapies and identification of biomarkers for the selection of patients with breast cancer for the bone targeted therapy in adjuvant setting

Bone metastases are responsible for different clinical complications defined as skeletal-related events (SREs) such as pathologic fractures, spinal cord compression, hypercalcaemia, bone marrow infiltration and severe bone pain requiring palliative radiotherapy. The general aim of these three years research period was to improve the management of patients with bone metastases through two different approaches of translational research. Firstly in vitro preclinical tests were conducted on breast cancer cells and on indirect co-colture of cancer cells and osteoclasts to evaluate bone targeted therapy singly and in combination with conventional chemotherapy. The study suggests that zoledronic acid has an antitumor activity in breast cancer cell lines. Its mechanism of action involves the decrease of RAS and RHO, as in osteoclasts. Repeated treatment enhances antitumor activity compared to non-repeated treatment. Furthermore the combination Zoledronic Acid + Cisplatin induced a high antitumoral activity in the two triple-negative lines MDA-MB-231 and BRC-230. The p21, pMAPK and m-TOR pathways were regulated by this combined treatment, particularly at lower Cisplatin doses. A co-colture system to test the activity of bone-targeted molecules on monocytes-breast conditioned by breast cancer cells was also developed. Another important criticism of the treatment of breast cancer patients, is the selection of patients who will benefit of bone targeted therapy in the adjuvant setting. A retrospective case-control study on breast cancer patients to find new predictive markers of bone metastases in the primary tumors was performed. Eight markers were evaluated and TFF1 and CXCR4 were found to discriminate between patients with relapse to bone respect to patients with no evidence of disease. In particular TFF1 was the most accurate marker reaching a sensitivity of 63% and a specificity of 79%. This marker could be a useful tool for clinicians to select patients who could benefit for bone targeted therapy in adjuvant setting.

Bone-implant interface. Evaluation of osteoblastic cells behavior on nanopatterned titanium surfaces: an in vitro analysis

Protein-adsorption occurs immediately following implantation of biomaterials. It is unknown at which extent protein-adsorption impacts the cellular events at bone-implant interface. To investigate this question, we compared the in-vitro outcome of osteoblastic cells grown onto titanium substrates and glass as control, by modulating the exposure to serum-derived proteins. Substrates consisted of 1) polished titanium disks; 2) polished disks nanotextured with H2SO4/H2O2; 3) glass. In the pre-adsorption phase, substrates were treated for 1h with αMEM alone (M-noFBS) or supplemented with 10%-foetal-bovine-serum (M-FBS). MC3T3-osteoblastic-cells were cultured on the pre-treated substrates for 3h and 24h, in M-noFBS and M-FBS. Subsequently, the culture medium was replaced with M-FBS and cultures maintained for 3 and 7days. Cell-number was evaluated by: Alamar-Blue and MTT assay. Mitotic- and osteogenic-activities were evaluated through fluorescence-optical-microscope by immunolabeling for Ki-67 nuclear-protein and Osteopontin. Cellular morphology was evaluated by SEM-imaging. Data were statistically analyzed using ANOVA-test, (p<0.05). At day3 and day7, the presence or absence of serum-derived proteins during the pre-adsorption phase had not significant effect on cell-number. Only the absence of FBS during 24h of culture significantly affected cell-number (p<0.0001). Titanium surfaces performed better than glass, (p<0.01). The growth rate of cells between day3 and 7 was not affected by the initial absence of FBS. Immunolabeling for Ki-67 and Osteopontin showed that the mitotic- and osteogenic- activity were ongoing at 72h. SEM-analysis revealed that the absence of FBS had no major influence on cell-shape. • Physico-chemical interactions without mediation by proteins are sufficient to sustain the initial phase of culture and guide osteogenic-cells toward differentiation. • The challenge is avoiding adsorption of ‘undesirables’ molecules that negatively impact on the cueing cells receive from surface. This may not be a problem in healthy patients, but may have an important role in medically-compromised-individuals in whom the composition of tissue-fluids is altered.

The biological basis of autism spectrum disorders: evaluation of oxidative stress and erytrocyte membrane alterations

This case-control study involved a total of 29 autistic children (Au) aged 6 to 12 years, and 28 gender and age-matched typically developing children (TD). We evaluated a high number of peripheral oxidative stress parameters, erythrocyte and lymphocyte membrane functional features and membrane lipid composition of erythrocyte. Erythrocyte TBARS, Peroxiredoxin II, Protein Carbonyl Groups and urinary HEL and isoprostane levels were elevated in AU (confirming an imbalance of the redox status of Au); other oxidative stress markers or associated parameters (urinary 8-oxo-dG, plasma Total antioxidant capacity and plasma carbonyl groups, erythrocyte SOD and catalase activities) were unchanged, whilst peroxiredoxin I showed a trend of elevated levels in red blood cells of Au children. A very significant reduction of both erythrocyte and lymphocyte Na+, K+-ATPase activity (NKA), a reduction of erythrocyte membrane fluidity, a reduction of phospatydyl serine exposition on erythrocyte membranes, an alteration in erythrocyte fatty acid membrane profile (increase in MUFA and in ω6/ω3 ratio due to decrease in EPA and DHA) and a reduction of cholesterol content of erythrocyte membrane were found in Au compared to TD, without change in erythrocyte membrane sialic acid content and in lymphocyte membrane fluidity. Some Au clinical features appear to be correlated with these findings; in particular, hyperactivity score appears to be related with some parameters of the lipidomic profile and membrane fluidity, and ADOS and CARS score are inversely related to peroxiredoxin II levels. Oxidative stress and erythrocyte structural and functional alterations may play a role in the pathogenesis of Autism Spectrum Disorders and could be potentially utilized as peripheral biomarkers.

A New Test Rig for In-Vitro Evaluation of the Knee Joint Behaviour

The evaluation of the knee joint behavior is fundamental in many applications, such as joint modeling, prosthesis and orthosis design. In-vitro tests are important in order to analyse knee behavior when simulating various loading conditions and studying physiology of the joint. A new test rig for in-vitro evaluation of the knee joint behavior is presented in this paper. It represents the evolution of a previously proposed rig, designed to overcome its principal limitations and to improve its performances. The design procedure and the adopted solution in order to satisfy the specifications are presented here. Thanks to its 6-6 Gough-Stewart parallel manipulator loading system, the rig replicates general loading conditions, like daily actions or clinical tests, on the specimen in a wide range of flexion angles. The restraining actions of knee muscles can be simulated when active actions are simulated. The joint motion in response to the applied loads, guided by passive articular structures and muscles, is permitted by the characteristics of the loading system which is force controlled. The new test rig guarantees visibility so that motion can be measured by an optoelectronic system. Furthermore, the control system of the new test rig allows the estimation of the contribution of the principal leg muscles in guaranteeing the equilibrium of the joint by the system for muscle simulation. Accuracy in positioning is guaranteed by the designed tibia and femur fixation systems,which allow unmounting and remounting the specimen in the same pose. The test rig presented in this paper permits the analysis of the behavior of the knee joint and comparative analysis on the same specimen before and after surgery, in a way to assess the goodness of prostheses or surgical treatments.

MRI-based breast volumetry-evaluation of three different software solutions

As lipofilling of the female breast is becoming more popular in plastic surgery, the use of MRI to assess breast volume has been employed to control postoperative results. Therefore, we sought to evaluate the accuracy of magnetic resonance imaging (MRI)-based breast volumetry software tools by comparing the measurements of silicone implant augmented breasts with the actual implant volume specified by the manufacturer. MRI-based volume analysis was performed in eight bilaterally augmented patients (46 ± 9 years) with three different software programs (Brainlab© I plan 2.6 neuronavigation software; mass analysis, version 5.3, Medis©; and OsiriX© v.3.0.2. 32-bit). The implant volumes analysed by the BrainLab© software had a mean deviation of 2.2 ± 1.7% (r?=?0.99) relative to the implanted prosthesis. OsiriX© software analysis resulted in a mean deviation of 2.8 ± 3.0% (r?=?0.99) and the Medis© software had a mean deviation of 3.1 ± 3.0% (r?=?0.99). Overall, the volumes of all analysed breast implants correlated very well with the real implant volumes. Processing time was 10 min per breast with each system and 30 s (OsiriX©) to 5 min (BrainLab© and Medis©) per silicone implant. MRI-based volumetry is a powerful tool to calculate both native breast and silicone implant volume in situ. All software solutions performed well and the measurements were close to the actual implant sizes. The use of MRI breast volumetry may be helpful in: (1) planning reconstructive and aesthetic surgery of asymmetric breasts, (2) calculating implant size in patients with missing documentation of a previously implanted device and (3) assessing post-operative results objectively.

Evaluation of perpendicular reflection intensity for assessment of caries lesion activity/inactivity

The aim of this study was to evaluate, using visual assessment, an experimental optical sensor measuring perpendicular reflection intensity (PRI) as an indicator of enamel caries lesion activity/inactivity. Forty teeth with either an active or an inactive enamel lesion were selected from a pool of extracted teeth. Each tooth was cut into halves, with a clinically sound half and a half with a non-cavitated enamel lesion. After gentle plaque removal, the teeth were kept moistened. The lesions were then photographed and a defined measuring site per lesion was chosen and indicated with an arrow on a printout. Independently, the chosen site was visually assessed for lesion activity, and its glossiness was measured with PRI assessment. Surface roughness (SR) was assessed with optical profilometry using a confocal microscope. Visual assessment and PRI were repeated after several weeks and a reliability analysis was performed. For enamel lesions visually scored as active versus inactive, significantly different values were obtained with both PRI and SR. PRI values of the clinically sound control surfaces were significantly different only from active lesions. Generally, inactive lesions had the same glossiness and the same roughness as the sound control surfaces. The reliabilities for visual assessment (? = 0.89) and for PRI (ICC = 0.86) were high. It is concluded that, within the limits of this study, PRI can be regarded as a promising tool for quantitative enamel lesion activity assessment. There is scope and potential for the PRI device to be considerably improved for in vivo use.

An Evaluation of Compression Schemes for Wireless Networks

Data gathering, either for event recognition or for monitoring applications is the primary intention for sensor network deployments. In many cases, data is acquired periodically and autonomously, and simply logged onto secondary storage (e.g. flash memory) either for delayed offline analysis or for on demand burst transfer. Moreover, operational data such as connectivity information, node and network state is typically kept as well. Naturally, measurement and/or connectivity logging comes at a cost. Space for doing so is limited. Finding a good representative model for the data and providing clever coding of information, thus data compression, may be a means to use the available space to its best. In this paper, we explore the design space for data compression for wireless sensor and mesh networks by profiling common, publicly available algorithms. Several goals such as a low overhead in terms of utilized memory and compression time as well as a decent compression ratio have to be well balanced in order to find a simple, yet effective compression scheme.

Evaluation of a novel needle guide for ultrasound-guided phantom vessel cannulation

We evaluated a novel, sled-mounted needle guide for ultrasound-guided vessel cannulation. Fifty medical students were randomly assigned to use ultrasound with the sled (sled group, n = 23) or ultrasound without the sled (control group, n = 27) for vessel cannulation in a phantom. For each of 15 attempts we recorded cannulation time and designated a successful cannulation as 1 and a failure as 0. Our primary outcome was the mean overall success rate. The median (IQR [range]) number of successes in the sled group and control group were 15.0 (13.0-15.0 [11.0-15.0]) and 11.0 (9.0-13.0 [6.0-15.0]), respectively (p < 0.001). Cannulation time decreased from the first to the last attempt in the sled group from 7.0 s (6.0-10.0 [4.0-16]) s to 4.0 s (3.0-4.0 [1.0-6.0]) s and in the control group from 35.0 s (27.0-35.0 [11.0-35.0]) s to 7.0 s (5.0-10.0 [3.0-25.0]) s. The sled group demonstrated a shorter cannulation time at each attempt (p < 0.001). The novel sled improved the success rate and efficiency of ultrasound-guided phantom vessel cannulation.

[Prophylaxis and therapy of postdural puncture headache--a critical evaluation of treatment options]

Since the first description of spinal and epidural anaesthesia, postdural puncture headache (PDPH) is a well known complication. Its prophylaxis and treatment has been studied and discussed for more than 100 years, but the evidence is still limited. Due to relatively low prevalence of PDPH, prospective RCTs are often missing, and the frequently self-limiting character of PDPH impedes an adequate interpretation of results from studies without a control group. Taking side effects and complications into account, a prophylactic treatment of PDPH cannot be recommended. In case of PDPH, non-opioid analgesics are the first choice treatment. The epidural blood patch remains the mainstay of severe PDPH therapy. Noninvasive therapies like theophylline, sumatriptan and ACTH can be an alternative. However, an evidence-based recommendation is lacking. The development of standard operating procedures for accidental dural punctures and PDPH is recommended.

Clinical and radiographic evaluation of intrabony periodontal defect treatment by open flap debridement alone or in combination with nanocrystalline hydroxyapatite bone substitute

The aim of this study has been to compare the clinical and radiographic outcome of periodontal intrabony defect treatment by open flap debridement alone or in combination with nanocrystalline hydroxyapatite bone substitute application. Thirty patients diagnosed with advanced periodontits were divided into two groups: the control group (OFD), in which an open flap debridement procedure was performed and the test group (OFD+NHA), in which defects were additionally filled with nanocrystalline hydroxyapatite bone substitute material. Plaque index (PI), gingival index (GI), bleeding on probing (BOP), pocket depth (PD), gingival recession (GR) and clinical attachment level (CAL) were measured prior to, then 6 and 12months following treatment. Radiographic depth and width of defects were also evaluated. There were no differences in any clinical and radiographic parameters between the examined groups prior to treatment. After treatment, BOP, GI, PD, CAL, radiographic depth and width parameter values improved statistically significantly in both groups. The PI value did not change, but the GR value increased significantly after treatment. There were no statistical differences in evaluated parameters between OFD and OFD+NHA groups 6 and 12months after treatment. Within the limits of the study, it can be concluded that the additional use of nanocrystalline hydroxyapatite bone substitute material after open flap procedure does not improve clinical and radiographic treatment outcome.

Evaluation of chemically modified SLA implants (modSLA) biofunctionalized with integrin (RGD)- and heparin (KRSR)-binding peptides

Enhancing osseointegration through surface immobilization of multiple short peptide sequences that mimic extracellular matrix (ECM) proteins, such as arginine-glycine-aspartic acid (RGD) and lysine-arginine-serine-arginine (KRSR), has not yet been extensively explored. Additionally, the effect of biofunctionalizing chemically modified sandblasted and acid-etched surfaces (modSLA) is unknown. The present study evaluated modSLA implant surfaces modified with RGD and KRSR for potentially enhanced effects on bone apposition and interfacial shear strength during early stages of bone regeneration. Two sets of experimental implants were placed in the maxillae of eight miniature pigs, known for their rapid wound healing kinetics: bone chamber implants creating two circular bone defects for histomorphometric analysis on one side and standard thread configuration implants for removal torque testing on the other side. Three different biofunctionalized modSLA surfaces using poly-L-lysine-graft-poly(ethylene glycol) (PLL-g-PEG) as a carrier minimizing nonspecific protein adsorption [(i) 20 pmol cm⁻² KRSR alone (KRSR); or in combination with RGD in two different concentrations; (ii) 0.05 pmol cm⁻² RGD (KRSR/RGD-1); (iii) 1.26 pmol cm⁻² RGD (KRSR/RGD-2)] were compared with (iv) control modSLA. Animals were sacrificed at 2 weeks. Removal torque values (701.48-780.28 N mm), bone-to-implant contact (BIC) (35.22%-41.49%), and new bone fill (28.58%-30.62%) demonstrated no significant differences among treatments. It may be concluded that biofunctionalizing modSLA surfaces with KRSR and RGD derivatives of PLL-g-PEG polymer does not increase BIC, bone fill, or interfacial shear strength.

Evaluation of smell and taste in patients with Wegener's granulomatosis

Although a reduced olfactory/gustatory function affects patients in all parts of life, this problem has not received much attention in Wegener's granulomatosis (WG). The aim of this study was to assess the smell/taste function of WG patients. Demographic data of 16 WG patients (9 males, 7 females) were obtained. They all subjectively assessed their taste/smell function on visual analogue scale. Olfactory/gustatory functions of the patients were tested with 'Sniffin' Sticks and 'Taste' strips, respectively. The results were then compared with those from sex and age-matched control group (n = 16) and normative data. WG patients subjectively assessed their olfactory (p = 0.03) and gustatory (p = 0.02) function to be lower than control group. All the olfactory scores (odour identification, odour discrimination and threshold) in both genders were significantly below the scores in the control group. WG patients were hyposmic. For taste (total taste score, as well as scores for the qualities sweet, sour, salty and bitter), WG patients did not significantly differ from controls and were normogeusic. However, the gustatory scores showed the tendency of reduction as compared to the control group. In conclusion, WG patients truly suffer from olfactory/taste dysfunction, but this is worse with olfaction. It is, therefore, imperative that physicians should make their patients to be aware of these sensory dysfunctions and educate them on methods to cope with it for better quality of life.

The evaluation of the oxidative state of low-density lipoproteins in intrauterine growth restriction and preeclampsia

To evaluate the oxidative state of lipoproteins in pregnancies complicated by intrauterine growth restriction (IUGR) in comparison to preeclampsia (PE) and healthy pregnant control subjects (CN).

Histomorphologic evaluation of extracorporeal shock wave therapy of the fourth metatarsal bone and the origin of the suspensory ligament in horses without lameness

OBJECTIVE: To determine via histologic examination and scintigraphy the effect of focused extracorporeal shock wave therapy (ESWT) on normal bone and the bone-ligament interface in horses. ANIMALS: 6 horses without lameness. PROCEDURE: Origins of the suspensory ligament at the metacarpus (35-mm probe depth) and fourth metatarsal bone (5-mm probe depth) were treated twice (days 0 and 16) with 2,000 shocks (energy flux density, 0.15 mJ/mm2). One forelimb and 1 hind limb were randomly treated, and the contralateral limbs served as nontreated controls. Bone scans were performed on days -1 (before ESWT), 3, 16, and 19. Histomorphologic studies of control and treated tissues were performed on day 30. RESULTS: ESWT significantly increased the number of osteoblasts but caused no damage to associated soft tissue structures and did not induce cortical microfractures. A significant correlation between osteoblast numbers and radiopharmaceutical uptake was noticed on lateral views of the hind limb on days 3 and 16 and on caudal views of the forelimb on day 3. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that ESWT has the potential to increase osteoblast numbers in horses. The correlation between increased osteoblast numbers and radio-pharmaceutical uptake 3 days and 16 days after the first ESWT suggested that stimulation of osteogenesis occurred soon after ESWT. No damage to bone or the bone-ligament interface should occur at the settings used in this study, and ESWT can therefore be administered safely in horses.

Serological and DNA-based evaluation of Chlamydia pneumoniae infection in inflammatory bowel disease

OBJECTIVES: Chlamydia has been associated with autoimmune diseases, but a link between chlamydial infection and the aetiopathogenesis of inflammatory bowel disease (IBD) remains controversial. In this study we assessed the relationship between chlamydial infection and IBD, as evidenced by serological measurement and DNA analysis of mucosal biopsy specimens. PATIENTS AND METHODS: The sera of 78 patients with Crohn's disease (CD), 24 patients with ulcerative colitis (UC), 73 healthy family members, and 20 healthy controls were tested for anti-C. pneumoniae IgG titres. A subgroup consisting of 13 UC and 39 CD patients was screened for the presence of chlamydial DNA on 42 inflamed versus 30 non-inflamed biopsy specimens and for mutations of their NOD2/CARD15 gene. RESULTS: Anti-C. pneumoniae IgG antibodies were found in the sera of 32 (41%) patients with CD, 11 (46%) patients with UC, 35 (48%) of unaffected family members, and nine (45%) unrelated healthy controls. Thirty-five percent of the control, 18% CD and 24% UC biopsy specimens contained C. pneumoniae DNA. In CD, however, C. pneumoniae DNA was significantly more frequently found in inflamed (27%) versus non-inflamed (8%) biopsy specimens (P < 0.05, Fisher's exact test). The frequencies of NOD2/CARD15 mutations were 33% for CD patients with C. pneumoniae DNA compared to 47% for CD patients without C. pneumoniae DNA. CONCLUSION: We found no marked differences in respect to anti-C. pneumoniae serum IgG or C. pneumoniae DNA between healthy controls and patients with IBD. However, in CD patients, inflamed tissue specimens contained significantly more likely C. pneumoniae DNA compared with biopsies from unaffected areas. Thus C. pneumoniae is unlikely to be of pathogenic importance in IBD while it may still influence local clinical manifestations.