Whole Genome Expression Profiling of Semitendinosus Tendons from Children with Diplegic and Tetraplegic Cerebral Palsy.

Cerebral palsy (CP) is the most common movement disorder in children, with a prevalence ranging from 1.5 to 4 per 1000 live births. CP is caused by a non-progressive lesion of the developing brain, leading to progressive alterations of the musculoskeletal system, including spasticity, often leading to the development of fixed contractures, necessitating tendon lengthening surgery. Total RNA-sequencing analysis was performed on semitendinosus tendons from diplegic and tetraplegic CP patients subjected to tendon lengthening surgery compared to control patients undergoing anterior cruciate ligament reconstructive surgery. Tetraplegic CP patients showed increased expression of genes implicated in collagen synthesis and extracellular matrix (ECM) turnover, while only minor changes were observed in diplegic CP patients. In addition, tendons from tetraplegic CP patients showed an enrichment for upregulated genes involved in vesicle-mediated transport and downregulated genes involved in cytokine and apoptotic signaling. Overall, our results indicate increased ECM turnover with increased net synthesis of collagen in tetraplegic CP patients without activation of inflammatory and apoptotic pathways, similar to observations in athletes where ECM remodeling results in increased tendon stiffness and tensile strength. Nevertheless, the resulting increased tendon stiffness is an important issue in clinical practice, where surgery is often required to restore joint mobility.

Antioxidants in smokers.

Cigarette smoke (CS) is likely the most common preventable cause of human morbidity and mortality worldwide. Consequently, inexpensive interventional strategies for preventing CS-related diseases would positively impact health systems. Inhaled CS is a powerful inflammatory stimulus and produces a shift in the normal balance between antioxidants and oxidants, inducing oxidative stress in both the respiratory system and throughout the body. This enduring and systemic pro-oxidative state within the body is reflected by increased levels of oxidative stress and inflammation biomarkers seen in smokers. Smokers might benefit from consuming antioxidant supplements, or a diet rich in fruit and vegetables, which can reduce the CS-related oxidative stress. This review provides an overview of the plasma profile of antioxidants observable in smokers and examines the heterogeneous literature to elucidate and discuss the effectiveness of interventional strategies based on antioxidant supplements or an antioxidant-rich diet to improve the health of smokers. An antioxidant-rich diet can provide an easy-to-implement and cost-effective preventative strategy to reduce the risk of CS-related diseases, thus being one of the simplest ways for smokers to stay in good health for as long as possible. The health benefits attributable to the intake of antioxidants have been observed predominantly when these have been consumed within their natural food matrices in an optimal antioxidant-rich diet, while these preventive effects are rarely achieved with the intake of individual antioxidants, even at high doses.

Guided Growth of the Proximal Femur for the Management of Hip Dysplasia in Children With Cerebral Palsy.

BACKGROUND: Progressive hip displacement is one of the most common and debilitating deformities seen in children with cerebral palsy (CP). The aim of this study was to evaluate the results of temporary medial hemiepiphysiodesis of the proximal femur (TMH-PF) using a transphyseal screw to control hip migration during growth in children with CP. METHODS: This was a retrospective study of children with CP and hip dysplasia, age 4 to 11 years and GMFCS levels III-V. There were 28 patients with 56 hips that underwent TMH-PF surgery between 2007 and 2010. Clinical and radiologic evaluation was performed preoperatively, at 6, 12, and 60 months following the index surgery. Acetabular index (AI), neck-shaft angle (NSA) and migration percentage (MP) were measured. All complications were recorded. RESULTS: All radiographic measurements were significantly improved at the final follow-up. Positive correlations were found between NSA, MP, and AI. Multiple regression analysis revealed that MP, time from surgery, and age were influenced by the decrease of the NSA. The femoral physis grew off the screw in 9 hips within 36 months. The screw head broke during attempted screw exchange in 1 hip. The remain cases (4 hips) were treated by placing a second screw parallel to the existing one. Finally, progressive subluxation occurred in 3 hips when the physis grew off the screw and were treated by skeletal reconstruction. CONCLUSIONS: TMH-PF was effective in controlling progressive subluxation of the hip in the majority of cases, obviating the need for major reconstructive surgery in these children with CP. LEVEL OF EVIDENCE: Level IV.

The Role of Taylor Spatial Frame in the Treatment of Blount Disease.

BACKGROUND: Alteration of the posteromedial part of the proximal tibia is the main characteristic of Blount's disease and if left untreated, leg alignment and normal development of the lower limbs may be compromised. AIM: To report treatment outcomes in children with Blount's disease using the Taylor Spatial Frame (TSF). MATERIALS AND METHODS: From January 2007 to December 2014, 16 young children (24 tibia) with a mean age of 7.5 years (range of 3-14 yrs) and severe Blount's disease were treated using TSF. Preoperative long standing radiographs were performed and anatomic medial proximal tibial angle (MPTA), diaphyseal-metaphyseal tibial angle (Drennan), femoro-tibial angle and leg length discrepancy (LLD) were measured. RESULTS: Post-operative improvement of all measurements was observed. MPTA increased from a mean of 71.8° (58° - 79°) to 92.5° (90° - 95°), the Drennan decreased from 16.6° (14° - 18°) to 3.6° (0° - 6°), the F-T angle changed from 15.4° (10° - 25°) of varus to 5.9° (2° - 10°) of valgus and the LLD decreased from 208 mm (150-320) to 69 mm (0- +120). Mean follow-up was 45.6 months. According to Paley's criteria pin track infection was present in 6 tibiae, while in 5 patients software changes were necessary. Recurrence was observed in 3 patients (triplets). Complete restoration of the mechanical axis was obtained at the end of the treatment. CONCLUSIONS: In the last decades, different surgical treatments have been proposed for Blount's disease (tension band plate, staples, osteotomies using external or internal fixation). External fixation using the TSF allows gradual safe correction of multiplanar deformities and is a well-tolerated technique by patients with Blount's disease.

Intra-Articular Cytokine Levels in Adolescent Patients after Anterior Cruciate Ligament Tear.

The treatment of anterior cruciate ligament (ACL) injuries in children and adolescents is challenging. Preclinical and clinical studies investigated ACL repairing techniques in skeletally immature subjects. However, intra-articular bioenvironment following ACL tear has not yet been defined in skeletally immature patients. The aim of this study was to measure cytokine concentrations in the synovial fluid in adolescent population. Synovial levels of IL-1ß, IL-1ra, IL-6, IL-8, IL-10, and TNF-α were measured in 17 adolescent patients (15 boys) with ACL tears who underwent ACL reconstruction including acute (5), subacute (7), and chronic (5) phases. Femoral growth plates were classified as "open" in three patients, "closing" in eight, and "closed" in six. Eleven patients presented an ACL tear associated with a meniscal tear. The mean Tegner and Lysholm scores (mean ± SD) of all patients were 8 ± 1 and 50.76 ± 26, respectively. IL-8, TNF-α, and IL-1ß levels were significantly greater in patients with "open" physes. IL-1ra and IL-1ß levels were significantly higher in patients with ACL tear associated with a meniscal tear. Poor Lysholm scores were associated with elevated IL-6 and IL-10 levels. IL-10 levels positively correlated with IL-6 and IL-8 levels, whereas TNF-α concentration negatively correlated with IL-6 levels. Skeletally immature patients with meniscal tears and open growth plates have a characteristic cytokine profile with particularly elevated levels of proinflammatory cytokines including IL-8, TNF-α, and IL-1ß. This picture suggests that the ACL tear could promote an intra-articular catabolic response in adolescent patients greater than that generally reported for adult subjects. The study lacks the comparison with synovial samples from healthy skeletally immature knees due to ethical reasons. Overall, these data contribute to a better knowledge of adolescent intra-articular bioenvironment following ACL injuries.

Assessment of glutathione/glutathione disulphide ratio and S-glutathionylated proteins in human blood, solid tissues, and cultured cells.

Glutathione (GSH) is the major non-protein thiol in humans and other mammals, which is present in millimolar concentrations within cells, but at much lower concentrations in the blood plasma. GSH and GSH-related enzymes act both to prevent oxidative damage and to detoxify electrophiles. Under oxidative stress, two GSH molecules become linked by a disulphide bridge to form glutathione disulphide (GSSG). Therefore, assessment of the GSH/GSSG ratio may provide an estimation of cellular redox metabolism. Current evidence resulting from studies in human blood, solid tissues, and cultured cells suggests that GSH also plays a prominent role in protein redox regulation via S -glutathionylation, i.e., the conjugation of GSH to reactive protein cysteine residues. A number of methodologies that enable quantitative analysis of GSH/GSSG ratio and S-glutathionylated proteins (PSSG), as well as identification and visualization of PSSG in tissue sections or cultured cells are currently available. Here, we have considered the main methodologies applied for GSH, GSSG and PSSG detection in biological samples. This review paper provides an up-to-date critical overview of the application of the most relevant analytical, morphological, and proteomics approaches to detect and analyse GSH, GSSG and PSSG in mammalian samples as well as discusses their current limitations.

Protein Carbonylation in Human Smokers and Mammalian Models of Exposure to Cigarette Smoke: Focus on Redox Proteomic Studies.

SIGNIFICANCE: Oxidative stress is one mechanism whereby tobacco smoking affects human health, as reflected by increased levels of several biomarkers of oxidative stress/damage isolated from tissues and biological fluids of active and passive smokers. Many investigations of cigarette smoke (CS)-induced oxidative stress/damage have been carried out in mammalian animal and cellular models of exposure to CS. Animal models allow the investigation of many parameters that are similar to those measured in human smokers. In vitro cell models may provide new information on molecular and functional differences between cells of smokers and nonsmokers. Recent Advances: Over the past decade or so, a growing number of researches highlighted that CS induces protein carbonylation in different tissues and body fluids of smokers as well as in in vivo and in vitro models of exposure to CS. CRITICAL ISSUES: We review recent findings on protein carbonylation in smokers and models thereof, focusing on redox proteomic studies. We also discuss the relevance and limitations of these models of exposure to CS and critically assess the congruence between the smoker's condition and laboratory models. FUTURE DIRECTIONS: The identification of protein targets is crucial for understanding the mechanism(s) by which carbonylated proteins accumulate and potentially affect cellular functions. Recent progress in redox proteomics allows the enrichment, identification, and characterization of specific oxidative protein modifications, including carbonylation. Therefore, redox proteomics can be a powerful tool to gain new insights into the onset and/or progression of CS-related diseases and to develop strategies to prevent and/or treat them. Antioxid. Redox Signal. 26, 406-426.

Knee arthrodesis by the Ilizarov method in the treatment of total knee arthroplasty failure.

BACKGROUND: Currently, the main indication for knee arthrodesis is septic failure of a total knee arthroplasty (TKA). The purpose of this study was to evaluate the results of knee arthrodesis by circular external fixation performed in the treatment of TKA failure in which revision arthroplasty was not indicated. METHODS: The study involved 19 patients who underwent knee arthrodesis by the Ilizarov method. Clinical and functional assessments were performed, including Knee Society Score (KSS). A postoperative clinical and radiographic evaluation was conducted every three months until the end of the treatment. Postoperative complications and eventual leg shortening were recorded. RESULTS: KSS results showed a significant improvement with respect to the preoperative condition. Of the 16 patients in the final follow-up, 15 patients (93.7%) achieved complete bone fusion. Major complications occurred in patients treated for septic failure of TKA and most occurred in patients over 75years of age; the mean final leg shortening was four centimeters. CONCLUSION: In our experience, the Ilizarov method is effective for performing a knee arthrodesis in the case of extensive bone loss. At the same time, it is possible to correct the associated leg deformities or limb length difference. In addition, only the Ilizarov method provides a mechanical stimulus for bone formation and improves the quality of the bone and of the microcirculation, which enhances the host response against infection. Despite these attributes, knee arthrodesis by the Ilizarov method must be considered a 'salvage procedure' in cases of severe outcomes from knee surgery in which revision arthroplasty is not indicated.

Can we prevent hip dislocation in children with cerebral palsy? Effects of postural management.

BACKGROUND: Hip dislocation is common in children with cerebral palsy (CP). At birth they do not have musculoskeletal deformities but they develop over time due to the combined effects of the movement disorder and impaired gross motor function. Early detection and treatment of a hip at risk is needed to modify the natural of hip development in CP. AIM: The aim of this study was to determine the effect of postural management treatment on hip displacement's progression in children CP. DESIGN: Prospective comparative non-randomized study. SETTING: Rehabilitative outpatient unit. POPULATION: Fifty-one children with CP were studied; the treated group (N.=30) was compared to a control group (N.=21). METHODS: The treated group followed a two year's long combined treatment program consisting a neurodevelopment treatment (NDT) two times a week and a 5 hours daily siège moulé postural program. The control group underwent only NDT twice a week for two years. Hip radiographs were measured with the migration percentage (MP) method at baseline, at 1 and 2 years of follow-up. RESULTS: A significant difference has been observed in the MP (%) trend (P<0.001) between treatment and control groups. At 2 years, there was a marked worsening (MP from 23.0 to 37.7) in the control group, compared to the stability (from 28.8 to 26.8) in the treatment group. CONCLUSIONS: This study supports the evidence that conservative postural management of hip deformity is useful to prevent the natural progression of hip dislocation. CLINICAL REHABILITATION IMPACT: Hip radiographic follow up program together with NDT and postural management program is useful to modify the natural progression of hip dislocation in children with CP.

Botulinum Toxin Treatment for Limb Spasticity in Childhood Cerebral Palsy.

CP is the most common cause of chronic disability in childhood occurring in 2-2.5/1000 births. It is a severe disorder and a significant number of patients present cognitive delay and difficulty in walking. The use of botulinum toxin (BTX) has become a popular treatment for CP especially for spastic and dystonic muscles while avoiding deformity and pain. Moreover, the combination of physiotherapy, casting, orthotics and injection of BTX may delay or decrease the need for surgical intervention while reserving single-event, multi-level surgery for fixed musculotendinous contractures and bony deformities in older children. This report highlights the utility of BTX in the treatment of cerebral palsy in children. We include techniques for administration, side effects, and possible resistance as well as specific use in the upper and lower limbs muscles.

Prevalence of Associated Lesions in Anterior Cruciate Ligament Reconstruction: Correlation With Surgical Timing and With Patient Age, Sex, and Body Mass Index.

BACKGROUND: Knee instability resulting from anterior cruciate ligament (ACL) rupture is an important risk factor for the onset of meniscal tears and cartilage injuries. A delay of the ligament reconstruction further increases this risk. There is currently no agreement on the right time for surgical ACL reconstruction. PURPOSE: To verify the correlation of time to ACL reconstruction, patient age, sex, and body mass index (BMI) with the prevalence of meniscal tears and cartilage injuries, as well as to identify the proper surgical timing to decrease the risk of developing associated injuries. STUDY DESIGN: Cohort study; Level of evidence, 3. METHODS: The medical records of 988 patients who underwent primary ACL reconstruction between January 2010 and May 2014 were analyzed to collect data on the prevalence of meniscal tears and cartilage injuries, surgical timing, and patient sex, age, and BMI. Logistic regression was performed to estimate the association between the prevalence of intra-articular lesions and the independent variables of surgical timing, sex, age, and BMI. RESULTS: The risk of developing at least an associated lesion increased by an average of 0.6% for each month of delay of surgical reconstruction. The odds ratio (OR) for developing an intra-articular lesion was 1.989 (95% CI, 1.403-2.820) in those waiting more than 12 months for ACL reconstruction. A 12-month delay for the intervention nearly doubled the risk of developing a medial meniscal tear (OR, 1.806 [95% CI, 1.317-2.475]) but did not modify the risk for the lateral meniscus (OR, 1.183 [95% CI, 0.847-1.653]). Concerning cartilage lesions, the risk after a 12-month delay increased in the medial compartment (femoral condyle: OR, 2.347 [95% CI, 1.499-3.676]; tibial plateau: OR, 5.574 [95% CI, 1.911-16.258]). In the lateral femoral condyle, the risk became significant in patients who underwent surgery more than 60 months after ACL injury as compared with those treated in the first 3 months (OR, 5.949 [95% CI, 1.825-19.385]). Lateral tibial plateau lesions did not seem to increase significantly. Male sex was a risk factor for the onset of lateral meniscal tears (OR, 2.288 [95% CI, 1.596-3.280]) and medial tears (OR, 1.752 [95% CI, 1.280-2.399]). Older age (OR, 1.017 [95% CI, 1.006-1.029]) and increased BMI (OR, 1.120 [95% CI, 1.072-1.169]) were risk factors for the occurrence of at least 1 associated lesion. CONCLUSION: ACL reconstruction within 12 months of injury can significantly reduce the risk of meniscal tears and chondral lesions. The close association between BMI and prevalence of associated lesions suggests that attention be paid to patients with an elevated BMI when considering the timing of ACL reconstruction surgery.

Plasma protein thiolation index (PTI) as a biomarker of thiol-specific oxidative stress in haemodialyzed patients.

The role of oxidative stress in patients with end stage renal disease (ESRD), which occurs at significantly higher levels than in the general population, is often underestimated in clinical practice. Emerging evidence highlights the strong correlation of oxidative stress with chronic inflammation and cardiovascular disease, which are highly prevalent in most patients on maintenance haemodialysis (HD) and are a major risk factor for mortality in this population. In this study, total plasma thiols and plasma S-thiolated proteins were measured in patients with ESRD, before and after a regular HD session, and compared to age-matched healthy subjects. We found a significant decrease in the level of total plasma thiols and, conversely, a significant increase in the level of S-thiolated proteins in these patients. In most patients, post-HD plasma level of total thiols did not differ from the one in healthy subjects, whereas plasma level of S-thiolated proteins was lower in HD patients than in age-matched healthy controls. This suggests that a single HD session restores plasma thiol redox status and re-establishes the antioxidant capacity of plasma thiols. Additionally, we determined protein thiolation index (PTI), i.e., the molar ratio between the sum of all low molecular mass thiols bound to S-thiolated plasma proteins and protein free cysteinyl residues. Patients with ESRD had a significantly higher PTI compared to age-matched healthy subjects and HD was associated with a decrease in PTI to normal, or lower than normal, levels. Although this study is limited in size, our results suggest that PTI is a useful indicator of thiol-specific oxidative stress in patients with ESRD on maintenance HD. This study also emphasizes that PTI determination is a cheap and simple tool suitable for large-scale clinical studies that could be used for routine screening of thiol-specific oxidative stress.

Intra-observer and inter-observer errors in CT measurement of torsional profiles of lower limbs: a retrospective comparative study.

BACKGROUND: The purpose of this study was to determine errors in measurement of torsional profiles (TP) (torsional femoral angle, torsional tibial angle, and femoral ankle angle) among four orthopedic surgeons, experts, and non-experts in measurement, and the learning curve. METHODS: Twenty-six lower extremities of 13 patients with spastic diplegia candidates for femoral/tibial derotational osteotomy had preoperative bilateral computer tomography (CT) scan grams to establish the TP. Each measurement was done by four orthopedic surgeons, two experienced clinicians and interpreters of CT imaging and two with limited clinical and imaging assessment experiences. Images were blinded and the surgeons made three determinations at least 5 days apart; the three angles were measured each time for each limb. Intra-observer and inter-observer variability were determined using bias, standard deviation, and interclass correlation coefficient. RESULTS: Significant inter-observer variability and bias were noted between experts and non-experts (average variability: ICC experts: 0.88 ± 0.15; ICC non-experts: 0.91 ± 0.09). For non-experts, excessive bias (25° and 14°) was observed. An associated improvement in bias with additional measurement experience indicated a potential significant learning curve for interpreting these studies. Less inter-observer variability was observed between experts. CONCLUSIONS: Measurement of TP is a reliable tool when used by experienced personnel, and their use as a preoperative tool should be reserved to ones with experience with such image assessments. Non-experts' measurements produced a weak agreement when compared to experts'.

I.S.Mu.L.T - Rotator Cuff Tears Guidelines.

Despite the high level achieved in the field of shoulder surgery, a global consensus on rotator cuff tears management is lacking. This work is divided into two main sessions: in the first, we set questions about hot topics involved in the rotator cuff tears, from the etiopathogenesis to the surgical treatment. In the second, we answered these questions by mentioning Evidence Based Medicine. The aim of the present work is to provide easily accessible guidelines: they could be considered as recommendations for a good clinical practice developed through a process of systematic review of the literature and expert opinion, in order to improve the quality of care and rationalize the use of resources.

Cigarette smoke induces alterations in the drug-binding properties of human serum albumin.

Albumin is the most abundant plasma protein and serves as a transport and depot protein for numerous endogenous and exogenous compounds. Earlier we had shown that cigarette smoke induces carbonylation of human serum albumin (HSA) and alters its redox state. Here, the effect of whole-phase cigarette smoke on HSA ligand-binding properties was evaluated by equilibrium dialysis and size-exclusion HPLC or tryptophan fluorescence. The binding of salicylic acid and naproxen to cigarette smoke-oxidized HSA resulted to be impaired, unlike that of curcumin and genistein, chosen as representative ligands. Binding of the hydrophobic fluorescent probe 4,4'-bis(1-anilino-8-naphtalenesulfonic acid) (bis-ANS), intrinsic tryptophan fluorescence, and susceptibility to enzymatic proteolysis revealed slight changes in albumin conformation. These findings suggest that cigarette smoke-induced modifications of HSA may affect the binding, transport and bioavailability of specific ligands in smokers.

Pathophysiology of tobacco smoke exposure: recent insights from comparative and redox proteomics.

First-hand and second-hand tobacco smoke are causally linked to a huge number of deaths and are responsible for a broad spectrum of pathologies such as cancer, cardiovascular, respiratory, and eye diseases as well as adverse effects on female reproductive function. Cigarette smoke is a complex mixture of thousands of different chemical species, which exert their negative effects on macromolecules and biochemical pathways, both directly and indirectly. Many compounds can act as oxidants, pro-inflammatory agents, carcinogens, or a combination of these. The redox behavior of cigarette smoke has many implications for smoke related diseases. Reactive oxygen and nitrogen species (both radicals and non-radicals), reactive carbonyl compounds, and other species may induce oxidative damage in almost all the biological macromolecules, compromising their structure and/or function. Different quantitative and redox proteomic approaches have been applied in vitro and in vivo to evaluate, respectively, changes in protein expression and specific oxidative protein modifications induced by exposure to cigarette smoke and are overviewed in this review. Many gel-based and gel-free proteomic techniques have already been used successfully to obtain clues about smoke effects on different proteins in cell cultures, animal models, and humans. The further implementation with other sensitive screening techniques could be useful to integrate the comprehension of cigarette smoke effects on human health. In particular, the redox proteomic approach may also help identify biomarkers of exposure to tobacco smoke useful for preventing these effects or potentially predictive of the onset and/or progression of smoking-induced diseases as well as potential targets for therapeutic strategies.

New insights in extracellular matrix remodeling and collagen turnover related pathways in cultured human tenocytes after ciprofloxacin administration.

We characterized the effect of ciprofloxacin (CPX) in cultured human tenocytes by morphological and molecular methods. Collagen type I and III mRNA and protein levels were unaffected, but lysyl hydroxylase 2b mRNA levels progressively decreased after CPX administration. MMP-1 protein levels significantly increased after 20 µg/ml CPX administration but remained unmodified at the higher dose, whilst MMP-2 activity was unchanged. Tissue inhibitor of MMP (TIMP-1) gene expression decreased after CPX treatment, whilst TIMP-2 and transforming growth factor-ß1 gene expression, the cytoskeleton arrangement, and cytochrome c expression remained unmodified. Secreted Protein Acidic and Rich in Cysteine mRNA and protein levels remained almost unchanged, whilst N-cadherin mRNA levels resulted significantly down-regulated and connexin 43 gene expression tended to decrease after CPX administration. The CPX-induced decreased ability to cross-link collagen and decreased TIMP-1 levels, possibly leading to higher activity of MMPs in ECM degradation, together with the down-regulation of N-cadherin and connexin 43 are consistent with a reduced ability to maintain tissue homeostasis, possibly making the tendon more susceptible to rupture.

Medium-term outcome in patients treated with total hip arthroplasty using a modular femoral stem.

The clinical, radiographic and quality of life results of total hip arthroplasty using the MODULUS cementless modular femoral stem were reviewed. 48 patients who had a total hip arthroplasty using the MODULUS femoral stem were identified. Six had bilateral procedures, resulting in 60 hips with complete clinical and radiographic data. Mean age at implantation was 50 years (range 33 to 82). Mean follow-up was 59 months (range 50 months to 73). There were two early post-operative dislocations (within 2 days). One patient required further surgery to remove heterotopic bone. Mean Harris Hip Score increased from 37 points preoperatively (range, 7 to 66) to 89 points at final review (range, 65 to 100 points). Radiographic evaluation revealed that all implants were stable without evidence of osteolysis but three patients (5%) exhibited heterotopic ossification. Quality of life was evaluated with the SF36. The physical component increased from 29.2 points (range, 18.5 to 46.0) to 51.7 points (range 42.9 to 60.6) and the mental component from 375 points (range, 19.5 to 50.0) to 50 points (range 32,8 to 62.0).

Pancreatic cancer cells retain the epithelial-related phenotype and modify mitotic spindle microtubules after the administration of ukrain in vitro.

The aim of this study is to characterize the phenotype of pancreatic ductal adenocarcinoma (PDAC) cells in relation to the expression of epithelial-to-mesenchymal transition (EMT) markers and determine whether ukrain, an anticancer drug based on the alkaloids extracted from greater celandine, modulates in vitro the malignant behavior of PDAC cells in order to extend our understanding of its therapeutic potential. Three cell lines (HPAF-II, HPAC, and PL45) were treated with ukrain (5, 10, and 20 µmol/l) for 48 h or left untreated (control). Cell proliferation was assessed by growth curves. Apoptosis was determined by Hoechst nuclear staining and by cytochrome c and caspase-8 expressions. The EMT markers E-cadherin, ß-catenin, and vimentin, as well as actin and tubulin cytoskeletons, were analyzed by immunofluorescence. Interphase and mitotic microtubules as well as abnormal mitotic figures were studied by fluorescence microscopy after tubulin immunolabeling. Ukrain strongly suppressed cell proliferation and induced apoptosis possibly through an extrinsic pathway as cytochrome c immunoreactivity suggested that the integrity of the mitochondria was not affected. Tubulin expression indicated an antiproliferative effect of ukrain on the basis of alterations in mitotic spindle microtubule dynamics, leading to abnormal mitosis. Membranous E-cadherin/ß-catenin immunoreactivity was similarly expressed in control-treated and ukrain-treated cells, although the drug upregulated E-cadherin in cell lysates. Our results suggest that ukrain exerts its chemotherapeutic action on PDAC cells targeting mitotic spindle microtubules, leading to abnormal mitosis and apoptosis, and favoring cell cohesiveness. The differentiated epithelial phenotype of HPAF-II, HPAC, and PL45 cell lines concomitant with a highly invasive potential suggests that further experiments will be necessary to definitively clarify the role of EMT in PDAC progression.

Red blood cells protect albumin from cigarette smoke-induced oxidation.

Different studies reported the presence of oxidized (carbonylated) albumin in the extravascular pool, but not in the intravascular one of cigarette smokers. In this study we attempted to explain this apparent discrepancy exposing human serum albumin (HSA) to aqueous cigarette smoke extract (CSE). CSE induces HSA carbonylation and oxidation of the HSA Cys34 sulfhydryl group. An antioxidant action of glutathione, cysteine, and its synthetic derivative N-acetylcysteine was observed only at supra-physiological concentrations, suggesting that physiological (plasma) concentrations of glutathione and cysteine in the low micromolar range are ineffective in preventing cigarette smoke-induced oxidation of HSA. Differently, human erythrocytes resulted to be protective towards CSE-induced oxidation (carbonylation and thiol oxidation) of both HSA and total human plasma proteins.