Screw Stress Distribution in a Clavicle Fracture with Plate Fixation: A Finite Element Analysis.

Clavicle midshaft fractures are mostly treated surgically by open internal reduction with a superior or anteroinferior plate and screws or by intramedullary nailing. Screw positioning plays a critical role in determining the stress distribution. There is a lack of data on the screw position and the appropriate number of cortices required for plate fixation. The aim of this study is to evaluate the mechanical behavior of an anterior plate implanted in a fractured bone subjected to 120° of lateral elevation compared to a healthy clavicle using numerical simulations. Contact forces and moments used were obtained from literature data and applied to the healthy and fractured finite element models. Stresses of about 9 MPa were found on the healthy clavicle, while values of about 15 MPa were calculated on the plate of the fractured one; these stress peaks were reached at about 30° and 70° of elevation when the stress shielding on the clavicle sums all the three components of the solicitation: compression, flexion, and torsion. The stress distribution in a clavicle fracture stabilized with plates and screws is influenced by several factors, including the plate's position and design, the type of screw, and the biomechanical forces applied during movements.

Association between neuropeptides and mucins in Crohn's disease mucous cells.

Crohn's disease (CD) and ulcerative colitis (UC) are both inflammatory bowel diseases (IBD). Unlike UC, which is limited to the mucosa of the colon, CD inflammation is characterized by chronic mucosal ulcerations affecting the entire gastrointestinal tract. Goblet cells (GCs) can be found in some lining epithelia, particularly in the respiratory and digestive tracts. GCs represent the main source of mucin that are the significant components of the mucus layer; hypertrophy of GCs and an increase in mucin production are observed in many enteric infections. The cytoplasm of goblet cells may also contain neuropeptides, such as serotonin, that can be altered in inflammatory bowel disease (IBD). The defense system of the gut is represented by the intestinal mucosal barrier, its protective function is strictly connected to the regulation of the mucus layer and the coordination of the neuro-immune response. Paraformaldehyde-fixed intestinal tissues, obtained from fifteen patients with Crohn's disease, were analyzed by immunostaining for MUC2, MUC4, 5-HT, and VAChT. This study aims to define the link between neuropeptides and mucins in mucous cells and their involvement in the inflammation process. Our results showed in mucous cells of Crohn's disease (CD) patients a high expression of MUC4 and a decrease in the expression of vesicular acetylcholine transporter (VAChT) demonstrating the presence of an inflammatory state.

The centripetal endoscopic sinus surgery in patients with cystic fibrosis: A preliminary study.

OBJECTIVES: The main aim of this study is to analyze the possible differences between clinical, demographic or genetic characteristics, in Cystic Fibrosis (CF) patients with chronic rhinosinusitis (CRS) with different phenotype. The secondary objective is to describe the possible benefit of surgery with Centripetal Endoscopic Sinus Surgery (CESS). METHODS: The study includes 56 who performed CT scan of the paranasal sinuses. They were divided in 3 group according to phenotype: CRS without Nasal Polyps (NP); CRS with NP; CRS complicated with Mucocele. The clinical symptoms, age, gender, genotype, microbial colonization and pulmonary disease stage were collected and analyzed to assess possible statistically significant differences. Regarding the 7 patients who performed CESS surgery, the number of hospitalizations, intravenous (iv) antibiotic courses, respiratory exacerbations, the FEV1, the Lund-Mackay Score (LMS) and the SNOT 22 were evaluated before and 1 year after surgery. RESULTS: No statistically significant differences regarding clinical symptoms between the 3 groups were identified (p > 0.05). Furthermore, there were no differences in age, gender, genotype, microbial colonization and pulmonary disease stage (p > 0.05). Regarding the patients who performed CESS, no significative difference in FEV1 progression was found. A reduction in hospitalization, pulmonary exacerbation and in the number of iv antibiotic courses resulted statistically significant different (p = 0.004; <0.001 and <0.001 respectively). A significant improvement in SNOT-22 and LMS (p < 0.001) was obtained. CONCLUSION: Radiological monitoring of the rhinosinus disease is necessary regardless of the clinical expression of the disease. The presence of CRS with NP complicated by mucocele is frequent and independent of the patient's age and clinical manifestations. An extensive surgical approach could represent the gold standard for patients with CF in consideration of the potential important advantages to perform a total toilet of all the sinuses and nasal cavities and at the same time eliminating a potential microbiological reservoir.

Vacuolated PAS-Positive Lymphocytes on Blood Smear: An Easy Screening Tool and a Possible Biomarker for Monitoring Therapeutic Responses in Late Onset Pompe Disease (LOPD).

Background: Primary aim was to investigate the diagnostic value of PAS-positive vacuolated lymphocytes on blood smear in Late Onset Pompe Disease (LOPD) patients and, secondly, to evaluate its potential utility in monitoring treatment effects. Methods: We examined blood smear of 26 LOPD patients. We evaluated 10 treated and 16 untreated LOPD patients. Among the latter group, 7 patients later initiated ERT and were tested again 6 months after start. Blood smear was also sampled from 82 controls and 19 patients with other muscle glycogenoses (MGSDs). PAS staining was used to evaluate: (1) presence of lymphocytes with glycogen-filled vacuoles, (2) quantification of vacuolated lymphocytes. Results: We found that PAS-positive lymphocytes were significantly higher in LOPD patients than in controls or other MGSDs (p < 0.05 and p < 0.001, respectively). ROC curve for discriminating between untreated LOPD patients and controls yielded an AUC of 1.00 (95%CI 1.00-1.00; p < 0.0001). PAS-positive lymphocyte cutoff level of >10 yielded sensitivity of 100% (95%CI 78-100%), specificity of 100% (95%CI 96-100%), and positive predictive value of 100%. Patients studied before and after ERT showed a dramatic decrease of PAS-positive vacuolated lymphocytes number (p = 0.016). In other MGSDs, PAS-positive lymphocytes were significantly lower that untreated LOPD patients but higher than controls. Conclusions: Our data suggest that the Blood Smear Examination (BSE) for PAS-positive lymphocytes quantification could be used as a simple and sensitive test for a quick screening of suspected Pompe disease. The quantification of vacuolated lymphocytes appears to be also a valuable tool for monitoring the efficacy of treatment in LOPD patients.

Ischemic stroke due to hypoperfusion in a patient with a previously unrecognized Danon disease.

Danon disease, an X-linked multisystemic disorder, is due to deficiency of Lysosome-Associated Membrane Protein 2 (LAMP2). It is usually characterized by hypertrophic cardiomyopathy, mental retardation and skeletal myopathy, sometimes also with atypical features. A 20-year-old man with cognitive impairment was admitted to the Emergency Room because of a sudden chest pain. ECG showed Wolff-Parkinson-White syndrome; echocardiography revealed hypertrophic cardiomyopathy, and, shortly after, he experienced a cardiac arrest followed by an occipital ischemic stroke. On neurological examination, he complained of visual loss, and diffuse muscle wasting and weakness were also unexpectedly noted. Electromyography evidenced a myopathic pattern and a peripheral neuropathy. A muscle biopsy disclosed vacuolar myopathy with glycogen storage; immunohistochemical studies demonstrated a LAMP-2 deficiency. LAMP2 molecular analysis identified a "de novo" mutation (p. Q353X). This patient with a neglected Danon disease, experienced an unusual complication as a stroke due to cerebral hypoperfusion after cardiac arrest caused by WPW syndrome.

The soy isoflavone genistein blunts nuclear factor kappa-B, MAPKs and TNF-α activation and ameliorates muscle function and morphology in mdx mice.

Several lines of evidence suggest a detrimental role of the nuclear factor-κB (NF-κB) activation in the dystrophic process. We showed in previous studies that its inhibition through drugs with antioxidant properties, have beneficial effects in mdx mice. We tested whether genistein, a well-known isoflavone, inhibitor of NF-κB, MAPK and TNF-α and readily available for clinical use, could have a beneficial effect in mdx mice in comparison with methylprednisolone, the gold standard treatment for DMD patients. Five-week old mdx mice received for five weeks: genistein (daily or 3-times/week), methylprednisolone or vehicle. Genistein treatment: (1) increased forelimb strength and strength normalized to weight; (2) reduced serum creatine-kinase levels; (3) reduced markers of oxidative stress; (4) reduced muscle necrosis and enhanced regeneration. The positive results were more evident with the daily administration of genistein and were comparable to the effect of corticosteroids. Our data support the novel hypothesis that, as other more specific therapeutic approaches are still under development, this soy-derived compound is a promising option to be further investigated in dystrophic process.

Flavocoxid counteracts muscle necrosis and improves functional properties in mdx mice: a comparison study with methylprednisolone.

Muscle degeneration in dystrophic muscle is exacerbated by the endogenous inflammatory response and increased oxidative stress. A key role is played by nuclear factor(NF)-kappaB. We showed that NF-kappaB inhibition through compounds with also antioxidant properties has beneficial effects in mdx mice, the murine model of Duchenne muscular dystrophy (DMD), but these drugs are not available for clinical studies. We evaluated whether flavocoxid, a mixed flavonoid extract with anti-inflammatory, antioxidant and NF-kappaB inhibiting properties, has beneficial effects in mdx mice in comparison with methylprednisolone, the gold standard treatment for DMD patients. Five-week-old mdx mice were treated for 5 weeks with flavocoxid, methylprednisolone or vehicle. The evaluation of in vivo and ex vivo functional properties and morphological parameters was performed. Serum samples were assayed for oxidative stress markers, creatine-kinase (CK) and leukotriene B-4. Cyclooxygenase-2 (COX-2), 5-lipoxygenase (5-LOX), tumor necrosis factor-alpha, p-38, JNK1 expression was evaluated in muscle by western blot analysis. NF-kappaB binding activity was investigated by electrophoresis mobility shift assay. The administration of flavocoxid: (1) ameliorated functional properties in vivo and ex vivo; (2) reduced CK; (3) reduced the expression of oxidative stress markers and of inflammatory mediators; (4) inhibited NF-kappaB and mitogen-activated protein kinases (MAPKs) signal pathways; (5) reduced muscle necrosis and enhanced regeneration. Our results highlight the detrimental effects of oxidative stress and NF-kappaB, MAPKs and COX/5-LOX pathways in the dystrophic process and show that flavocoxid is more effective in mdx mice than methylprednisolone.

Polydeoxyribonucleotide (PDRN) restores blood flow in an experimental model of peripheral artery occlusive disease.

OBJECTIVE: This study investigated whether polydeoxyribonucleotide (PDRN) may be efficacious in the treatment of peripheral artery occlusive diseases, which are a major cause of morbidity in Western countries and still lack standardized treatment. METHODS: We investigated the effects of PDRN, a mixture of deoxyribonucleotides, in an experimental model of hind limb ischemia (HLI) in rats to stimulate vascular endothelial growth factor (VEGF)-A production and to avoid critical ischemia. The femoral artery was excised to induce HLI. Sham-operated on rats (sham HLI) were used as controls. Animals were treated daily with intraperitoneal PDRN (8 mg/kg) or its vehicle. Animals were euthanized at day 7, 14, and 21 after the evaluation of blood flow by laser Doppler. Dissected muscles were used to measure VEGF-A messenger RNA (mRNA) and protein expression, to evaluate edema, and to assess histologic damage. RESULTS: Administration of PDRN dramatically increased VEGF mRNA throughout the study (day 14: HLI, 7 +/- 2.2 n-fold/beta-actin; HLI + PDRN, 13.3 +/- 3.8 n-fold/beta-actin; P < .0001) and protein expression (HLI, 11 +/- 3.4 integrated intensity; HLI + PDRN, 16 +/- 3.8 integrated intensity; P < .0001). The compound stimulated revascularization, as confirmed by blood flow restoration (P < .005 vs HLI + vehicle), and blunted the histologic damage and the degree of edema. PDRN did not modify VEGF-A expression and blood flow in sham HLI animals. Furthermore, the concomitant administration of 3,7-dimethyl-1-propargilxanthine (DMPX), a selective adenosine A(2A) receptor antagonist, abolished the positive effects of PDRN, confirming that PDRN acts through this receptor. CONCLUSION: These results led us to hypothesize a role for PDRN in treating peripheral artery occlusive diseases.

Simvastatin enhances VEGF production and ameliorates impaired wound healing in experimental diabetes.

Statins have different effects beyond cholesterol reduction and stimulate angiogenesis. We investigated the effect of simvastatin in diabetes-related healing defects. An incisional skin wound model produced on the back of female diabetic mice (db(+)/db(+)) and their normoglycemic littermates (db(+)/(+)m) was used. Animals were treated daily either with simvastatin (5 mg/(kgi.p.)) or vehicle. Mice were killed on different days (3, 6 and 12 after skin injury) for measurement of vascular endothelial growth factor (VEGF) mRNA and protein expression, to assess histologically the healing process and to evaluate wound breaking strength and angiogenesis by CD31 immunostaining. Simvastatin administration in diabetic mice increased VEGF mRNA (simvastatin=4.8+/-0.6n-fold/beta-actin; vehicle=2.3+/-0.4n-fold/beta-actin) and protein expression (simvastatin=5+/-0.7 integrated intensity; vehicle=2.2+/-0.3 integrated intensity) and enhanced nitric oxide wound content at day 6. Additionally, the statin augmented breaking strength and PECAM-1 immunostaining at day 12. Finally, simvastatin administration restored the impaired wound healing process in diabetic mice. Similar results were obtained in normoglycaemic mice. Passive immunization with anti-VEGF antibody (10 microg/mouse) completely abrogated the beneficial effects of simvastatin on healing in diabetic mice. Simvastatin has potential application in diabetes-related wound healing disorders.

VEGF overexpression via adeno-associated virus gene transfer promotes skeletal muscle regeneration and enhances muscle function in mdx mice.

Vascular endothelial growth factor (VEGF) is a major regulator of physiological and pathological angiogenesis. Recently it was reported that the delivery of VEGF using recombinant adeno-associated virus (rAAV) vectors reduces muscle damage and promotes muscle regeneration in different experimental models of muscle necrosis. We demonstrate that intramuscular administration of rAAV-VEGF improved pathophysiology of the mdx mouse, a model of Duchenne muscular dystrophy (DMD). One month after injection, rAAV-VEGF-treated muscles showed augmented expression of VEGF and immunolocalization of its receptor, VEGFR-2. VEGF-treated mdx mice showed increased forelimb strength and strength normalized to weight. Treatment reduced necrotic fibers area and increased regenerating fibers area with an augmented number of myogenin-positive satellite cells and myonuclei, and of developmental myosin heavy chain-positive fibers. Only the regenerating area showed increased capillary density. This study provides novel evidence of a VEGF beneficial effect in mdx mice that is exerted mainly by a proregenerative and angiogenic effect. It opens new therapeutic prospectives in DMD and other types of muscular disorders.

Immunohistochemical analysis of human skeletal muscle AMP deaminase deficiency. Evidence of a correlation between the muscle HPRG content and the level of the residual AMP deaminase activity.

We have previously described that, in healthy human skeletal muscle, an anti-histidine-proline-rich-glycoprotein (HPRG) antibody selectively binds to type IIB fibers that are well known to contain the highest level of AMP deaminase (AMPD) activity, suggesting an association of the HPRG-like protein to the enzyme isoform M. The present paper reports an immunohistochemical study performed on human skeletal muscle biopsies from patients with AMPD deficiency and carried out utilizing both the anti-HPRG antibody and an anti-AMPD antibody specific for the isoform M. A correlation between the muscle content of the HPRG-like protein and the level of AMPD activity was demonstrated. In the specimens from patients with Acquired AMPD deficiency the HPRG-immunoreactivity was less intense than that shown by the control subjects and was related to the residual AMPD activity. The patients affected by Primary and Coincidental AMPD deficiency, which were characterized by an absence of enzyme activity and AMPD immunoreactivity, showed the lowest HPRG immunoreactivity that was clearly detectable by Western blot analysis, but not by immunohistochemistry. The interpretation of the significance of these observations suggests a physiological mutual dependence between skeletal muscle HPRG and AMPD polypeptides with regard to their stability.

Asymptomatic hyperCKemia in a case of Danon disease due to a missense mutation in Lamp-2 gene.

Primary lysosome-associated membrane protein-2 (LAMP-2) deficiency is an X-linked disease, characterized by the clinical triad of cardiomyopathy, vacuolar myopathy and mental retardation, previously known as Danon disease. Mutations of lamp-2 gene have been reported so far in about 20 patients, one of whom was Italian. We describe a new Italian case with persistent hyperCKemia, exercise intolerance and hypertrophic cardiomyopathy but with no muscle weakness or mental impairment. Muscle biopsy revealed a vacuolar myopathy with mild glycogen storage, and immunohistochemical studies detected LAMP-2 deficiency. A new nucleotide substitution (T961C) on exon 8 of lamp-2 gene was identified as responsible for the protein deficiency. This is the first missense mutation so far described. LAMP-2 deficiency should be considered as a cause of recurrent hyperCKemia and hypertrophic cardiomyopathy.