Peroneus Brevis flap in Achilles tendon reconstruction. Clinical, radiological and functional analysis.

BACKGROUND: We would like to describe our experience with Peroneus Brevis flap in complicated Achilles tendon re-ruptures with fringed stumps. METHODS: Eight patients with monolateral re-rupture of Achilles tendon were selected as eligible for surgical repair with Peroneus Brevis flap. Patients' outcome was evaluated clinically (ATRS and ROM), functionally (Gait analysis) and MRI was performed before and after surgery. RESULTS: Effective coverage of tissue defect was reached in all patients. Functional assessment evaluation results were registered in a follow-up time that ranged from 12 to 18 months. ATRS and ROM tests' results showed good functional recovery without functional limitations or subjective reports pain. Post-operative MRI showed no signs of inflammation or tissue gaps. Gait analysis showed a partial reduction of performance in the affected side that did not affect patients' quality of life. CONCLUSIONS: In the presence of fringed stumps in Achilles tendon re-rupture, tendon flaps have the benefits of autologous tissues transfers and present less risks of failure than free flaps. Among them, Peroneus Brevis flap is easy to perform and leads to donor site's low morbidity. Our preliminary experience provides support for this technique to be potentially validated in larger more controlled trial.

The effectiveness of negative pressure therapy on infected wounds: preliminary results.

Vacuum-assisted closure (VAC) therapy is a sophisticated system that maintains a closed, humid, sterile and isolated environment. Wound infection is considered a relative contraindication. The objective of this study is to extend the indications for VAC therapy to include infected wounds by demonstrating its ability to increase the antibiotic concentration in the damaged and infected tissues. Patients who presented with ulcers infected with daptomycin-sensitive bacteria were eligible to be enrolled in this prospective study. They were given antibiotic therapy with daptomycin with a specific protocol. A biopsy of the lesion was carried out to detect tissue concentration of the drug at time 0. Afterwards, the patients were subjected to VAC therapy. At the end of VAC therapy, a second lesion biopsy was performed and analysed to detect tissue concentration of the drug at time 1. A control group was enrolled in which patients followed the same protocol, but they were treated with traditional dressings. Fisher's exact test was used to compare the two groups. The results highlighted a significant increase in the concentration of antibiotics in the study group tissue; the improvement was sensibly lower in the control group. Statistical differences were not found between the two groups. The preliminary analysis of the data showed an important increase of antibiotic concentration in the tissue after VAC therapy. Despite the encouraging data, it is necessary to broaden the sample of patients and perform the same study with other antibiotics.

Effects of oxaliplatin and oleic acid Gc-protein-derived macrophage-activating factor on murine and human microglia.

The biological properties and characteristics of microglia in rodents have been widely described, but little is known about these features in human microglia. Several murine microglial cell lines are used to investigate neurodegenerative and neuroinflammatory conditions; however, the extrapolation of the results to human conditions is frequently met with criticism because of the possibility of species-specific differences. This study compares the effects of oxaliplatin and of oleic acid Gc-protein-derived macrophage-activating factor (OA-GcMAF) on two microglial cell lines, murine BV-2 cells and human C13NJ cells. Cell viability, cAMP levels, microglial activation, and vascular endothelial growth factor (VEGF) expression were evaluated. Our data demonstrate that oxaliplatin induced a significant decrease in cell viability in BV-2 and in C13NJ cells and that this effect was not reversed with OA-GcMAF treatment. The signal transduction pathway involving cAMP/VEGF was activated after treatment with oxaliplatin and/or OA-GcMAF in both cell lines. OA-GcMAF induced a significant increase in microglia activation, as evidenced by the expression of the B7-2 protein, in BV-2 as well as in C13NJ cells that was not associated with a concomitant increase in cell number. Furthermore, the effects of oxaliplatin and OA-GcMAF on coculture morphology and apoptosis were evaluated. Oxaliplatin-induced cell damage and apoptosis were nearly completely reversed by OA-GcMAF treatment in both BV-2/SH-SY5Y and C13NJ/SH-SY5Y cocultures. Our data show that murine and human microglia share common signal transduction pathways and activation mechanisms, suggesting that the murine BV-2 cell line may represent an excellent model for studying human microglia.

Gc-protein-derived macrophage activating factor counteracts the neuronal damage induced by oxaliplatin.

Oxaliplatin-based regimens are effective in metastasized advanced cancers. However, a major limitation to their widespread use is represented by neurotoxicity that leads to peripheral neuropathy. In this study we evaluated the roles of a proven immunotherapeutic agent [Gc-protein-derived macrophage activating factor (GcMAF)] in preventing or decreasing oxaliplatin-induced neuronal damage and in modulating microglia activation following oxaliplatin-induced damage. The effects of oxaliplatin and of a commercially available formula of GcMAF [oleic acid-GcMAF (OA-GcMAF)] were studied in human neurons (SH-SY5Y cells) and in human microglial cells (C13NJ). Cell density, morphology and viability, as well as production of cAMP and expression of vascular endothelial growth factor (VEGF), markers of neuron regeneration [neuromodulin or growth associated protein-43 (Gap-43)] and markers of microglia activation [ionized calcium binding adaptor molecule 1 (Iba1) and B7-2], were determined. OA-GcMAF reverted the damage inflicted by oxaliplatin on human neurons and preserved their viability. The neuroprotective effect was accompanied by increased intracellular cAMP production, as well as by increased expression of VEGF and neuromodulin. OA-GcMAF did not revert the effects of oxaliplatin on microglial cell viability. However, it increased microglial activation following oxaliplatin-induced damage, resulting in an increased expression of the markers Iba1 and B7-2 without any concomitant increase in cell number. When neurons and microglial cells were co-cultured, the presence of OA-GcMAF significantly counteracted the toxic effects of oxaliplatin. Our results demonstrate that OA-GcMAF, already used in the immunotherapy of advanced cancers, may significantly contribute to neutralizing the neurotoxicity induced by oxaliplatin, at the same time possibly concurring to an integrated anticancer effect. The association between these two powerful anticancer molecules would probably produce the dual effect of reduction of oxaliplatin-induced neurotoxicity, together with possible synergism in the overall anticancer effect.

Analysis and Report of the Physical and Rehabilitation Medicine Evaluation Activity in Patients Admitted to Acute Care Setting: An Observational Retrospective Study.

BACKGROUND: Disability (both temporary and transitory, or definitive) might occur for the first time in a given patient after an acute clinical event. It is essential, whenever indicated, to undergo a Physical Medicine and Rehabilitation assessment to detect disability and any need for rehabilitation early. Although access to rehabilitation services varies from country to country, it should always be governed by a PRM prescription. OBJECTIVE: The aim of the present observational retrospective study is to describe consultancy activity performed by PRM specialists in a university hospital in terms of requests' typology, clinical questions, and rehabilitation setting assignment. METHODS: Multiple parameters were analyzed (clinical condition, patient's socio-family background, and rehabilitation assessment scale scores) and a correlation analysis was performed between the analyzed characteristics and both the different clinical conditions and the assigned rehabilitation setting. RESULTS: PRM evaluations of 583 patients from 1 May 2021 to 30 June 2022 were examined. Almost half of the total sample (47%) presented disability due to musculoskeletal conditions with a mean age of 76 years. The most frequently prescribed settings were home rehabilitation care, followed by intensive rehabilitation and long-term care rehabilitation. CONCLUSIONS: Our results suggest the high public health impact of musculoskeletal disorders, followed by neurological disorders. This is, however, without forgetting the importance of early rehabilitation to prevent other types of clinical conditions such as cardiovascular, respiratory, or internal diseases from leading to motor disability and increasing costs.

Tensegrity and plasma for skin regeneration.

BACKGROUND: Mechanical stresses induce variations in tissue tensegrity leading to cell proliferation and differentiation thus contributing to tissue remodelling. Besides mechanical forces, skin remodelling may be induced by the application of plasma, a new type of energy delivery resulting in controlled heat damage. Here we demonstrate that mechanical stress induced by the application of vacuum increases the efficacy of plasma in skin regeneration treatment. METHODS: Vacuum alone and vacuum plus plasma at different energies were applied to rat skin and biopsies collected at different time intervals after treatments. Skin integrity, collagen arrangement, inflammation and myofibroblast differentiation were assessed by Masson's trichrome staining. Procollagen synthesis was evaluated by immunohistochemistry. RESULTS: Vacuum alone induced significant and temporary alterations in the distribution of collagen bundles, with concomitant procollagen synthesis in the dermis; no myofibroblasts and no signs of inflammation were observed. Vacuum plus plasma determined an important spatial modification of collagen bundles, more intense than vacuum or plasma alone. Significant increase of procollagen synthesis, numerous myofibroblasts but slight sign of inflammation appeared after the treatment. CONCLUSION: Vacuum mechanically stimulated fibroblasts, producing changes in collagen arrangement and procollagen synthesis. Plasma led to the same effects through thermal damage. Application of a combined treatment consisting in vacuum plus plasma induced more remarkable effects on skin regeneration with relatively low plasma energies and no relevant side effects.

Non-Surgical and Rehabilitative Interventions in Patients with Frozen Shoulder: Umbrella Review of Systematic Reviews.

Background: Frozen shoulder (FS) is a painful condition characterized by progressive loss of shoulder function with passive and active range of motion reduction. To date, there is still no consensus regarding its rehabilitative treatment for pain management. Purpose: The aim of this umbrella review of systematic reviews was to analyze the literature, investigating the effects of non-surgical and rehabilitative interventions in patients suffering from FS. Patients and Methods: A review of the scientific literature was carried out from 2010 until April 2020 using the following search databases: PubMed, Medline, PEDro, Scopus and Cochrane Library of Systematic Reviews. A combination of terms was used for the search: frozen shoulder OR adhesive capsulitis AND systematic review OR meta-analysis AND rehabilitation NOT surgery NOT surgical intervention. We included systematic reviews that specifically dealt with adults with FS, treated with non-surgical approaches. All the systematic reviews and meta-analyses included in the study that met the inclusion criteria were assessed using the Assessment of Multiple Systematic Reviews as a quality assessment tool. Results: Out of 49 studies, only 14 systematic reviews respected the eligibility criteria and were included in this study. Their results showed an important heterogeneity of the studies and all of them agree on the lack of high-quality scientific work to prove unequivocally which rehabilitative treatment is better than the other. Due to this lack of gold standard criteria, there may be also a heterogeneity in the diagnosis of the reviews analyzed. Conclusion: Non-surgical and rehabilitative interventions are undoubtedly effective in treating FS, but there is no evidence that one approach is more effective than the other regarding the methods reported. Future high-quality RCTs are needed to standardize the treatment modalities of each physiotherapy intervention to provide strong recommendations in favor.

Gc protein-derived macrophage-activating factor (GcMAF) stimulates cAMP formation in human mononuclear cells and inhibits angiogenesis in chick embryo chorionallantoic membrane assay.

The effects of Gc protein-derived macrophage-activating factor (GcMAF) have been studied in cancer and other conditions where angiogenesis is deregulated. In this study, we demonstrate for the first time that the mitogenic response of human peripheral blood mononuclear cells (PBMCs) to GcMAF was associated with 3'-5'-cyclic adenosine monophosphate (cAMP) formation. The effect was dose dependent, and maximal stimulation was achieved using 0.1 ng/ml. Heparin inhibited the stimulatory effect of GcMAF on PBMCs. In addition, we demonstrate that GcMAF (1 ng/ml) inhibited prostaglandin E(1)- and human breast cancer cell-stimulated angiogenesis in chick embryo chorionallantoic membrane (CAM) assay. Finally, we tested different GcMAF preparations on CAM, and the assay proved to be a reliable, reproducible and inexpensive method to determine the relative potencies of different preparations and their stability; we observed that storage at room temperature for 15 days decreased GcMAF potency by about 50%. These data could prove useful for upcoming clinical trials on GcMAF.

Angiotensin-converting enzyme/vitamin D receptor gene polymorphisms and bioelectrical impedance analysis in predicting athletic performances of Italian young soccer players.

We evaluated the association between 2 genetic polymorphisms known to be involved in fitness and performance, and anthropometric features, body composition, and athletic performances in young male soccer players with the goal of identifying genetic profiles that can be used to achieve maximal results from training. One hundred twenty-five medium-high-level male soccer players were genotyped for angiotensin-converting enzyme (ACE) I/D, and vitamin D receptor (VDR) FokI gene polymorphisms and scored for anthropometric measurements, body composition, and athletic performance. Body mass index, fat mass, fat-free mass, resistance, reactance, impedance, phase angle (PA), and body cell mass were measured. Athletic performance was evaluated by squat jump, countermovement jump (CMJ), 2-kg medicine ball throw, 10- and 20-m sprint time. We observed that the homozygous ff genotype of the VDR gene was significantly more represented in young soccer players than in a matched sedentary population. Values of reactance and PA were differently distributed in ACE and VDR genotypes with high mean values in subjects with DD (ACE) and FF (VDR) genotypes. No correlation was observed between ACE or VDR genotypes and 2-kg medicine ball throw, 10- and 20-m sprint times. The ID genotype of ACE was associated with the best performances in squat jump and CMJ. Our results suggest that determination of ACE and VDR genotypes might help select those young athletes harboring the most favorable genetic potential to succeed in soccer.

State of the evidence about rehabilitation interventions in patients with dysphagia.

INTRODUCTION: Rather than a separate nosological entity, dysphagia must be considered as a symptom of other pathological conditions, which afflicts patients admitted to numerous medical departments (rehabilitation, neurology, geriatrics, internal medicine, etc.) These disorders share the need for timely access to quality care and multidisciplinary treatment, including rehabilitation. The purpose of this study was to conduct a review of the current guidelines' recommendations in the literature and provide recommendations on the rehabilitative management of the patient with dysphagia. EVIDENCE ACQUISITION: The search was carried out through the main databases (Medline, Pedro, Cochrane Database and Google Scholar). All the articles concerning rehabilitation management of dysphagia, published in the last 10 years, have been included. EVIDENCE SYNTHESIS: Bibliographic research has provided thirteen guidelines. The literature analysed focuses mainly on the screening, the evaluation and the planning of multidisciplinary treatment. The literature agrees in recommending as cornerstones in the treatment of the dysphagic patient dietary changes, rehabilitation training (particularly muscle strengthening exercises and coordination) and early use of alternative nutrition in patients severely compromised. CONCLUSIONS: The dysphagic patient requires the deployment of a range of skills by a multi-professional and multi-disciplinary team. Speech and language pathologists in cooperation with specialists of rehabilitation have the task of managing the various stages, ranging from the early identification of the symptom to the setting of the treatment plan. Due to the lack of standardized protocols, it is necessary to implement the research path, especially regarding rehabilitation intervention.

Motor Recovery After Stroke: From a Vespa Scooter Ride Over the Roman Sampietrini to Focal Muscle Vibration (fMV) Treatment. A 99mTc-HMPAO SPECT and Neurophysiological Case Study.

Focal repetitive muscle vibration (fMV) is a safe and well-tolerated non-invasive brain and peripheral stimulation (NIBS) technique, easy to perform at the bedside, and able to promote the post-stroke motor recovery through conditioning the stroke-related dysfunctional structures and pathways. Here we describe the concurrent cortical and spinal plasticity induced by fMV in a chronic stroke survivor, as assessed with 99mTc-HMPAO SPECT, peripheral nerve stimulation, and gait analysis. A 72-years-old patient was referred to our stroke clinic for a right leg hemiparesis and spasticity resulting from a previous (4 years before) hemorrhagic stroke. He reported a subjective improvement of his right leg's spasticity and dysesthesia that occurred after a30-min ride on a Vespa scooter as a passenger over the Roman Sampietrini (i.e., cubic-shaped cobblestones). Taking into account both the patient's anecdote and the current guidelines that recommend fMV for the treatment of post-stroke spasticity, we then decided to start fMV treatment. 12 fMV sessions (frequency 100 Hz; amplitude range 0.2-0.5 mm, three 10-min daily sessions per week for 4 consecutive weeks) were applied over the quadriceps femoris, triceps surae, and hamstring muscles through a specific commercial device (Cro®System, NEMOCOsrl). A standardized clinical and instrumental evaluation was performed before (T0) the first fMV session and after (T1) the last one. After fMV treatment, we observed a clinically relevant motor and functional improvement, as assessed by comparing the post-treatment changes in the score of the Fugl-Meyer assessment, the Motricity Index score, the gait analysis, and the Ashworth modified scale, with the respective minimal detectable change at the 95% confidence level (MDC95). Data from SPECT and peripheral nerve stimulation supported the evidence of a concurrent brain and spinal plasticity promoted by fMV treatment trough activity-dependent changes in cortical perfusion and motoneuron excitability, respectively. In conclusion, the substrate of post-stroke motor recovery induced by fMV involves a concurrently acting multisite plasticity (i.e., cortical and spinal plasticity). In our patient, operant conditioning of both cortical perfusion and motoneuron excitability throughout a month of fMV treatment was related to a clinically relevant improvement in his strength, step symmetry (with reduced limping), and spasticity.

Chronic kidney disease and vitamin D: how much is adequate?

Mehrotra et al. demonstrate that there still is hypovitaminosis D in adults with chronic kidney disease (CKD) in the United States, and this defect is associated with increased risk for death. Definition of the adequate amount of vitamin D, however, is still uncertain; polymorphisms of the gene encoding the vitamin D receptor might be responsible for this uncertainty. People carrying less efficient variants of the receptor might need higher amounts of vitamin D.

Replicating neuroblastoma cells in different cell cycle phases display different vulnerability to amyloid toxicity.

A key role of mitotic activation in neuronal cell death in early stages of Alzheimer's disease (AD) has been suggested. Apparently, terminally differentiated neurons are precluded from mitotic division, yet some phenotypic markers of cell cycling are present in AD-vulnerable brain areas. In this paper, we investigated whether dividing human neuroblastoma cells are preferentially vulnerable to amyloid aggregate toxicity in some specific cell cycle stage(s). Our data indicate that Abeta1-40/42 aggregates added to the cell culture media bind to the plasma membrane and are internalized faster in the S than in the G2/M and G1 cells possibly as a result of a lower content in membrane cholesterol in the former. Earlier and sharper increases in reactive oxygen species production triggered a membrane oxidative injury and a significant impairment of antioxidant capacity, eventually culminating with apoptotic activation in S and, to a lesser extent, in G2/M exposed cells. G1 cells appeared more resistant to the amyloid-induced oxidative attack possibly because of their higher antioxidant capacity. The high vulnerability of S cells to aggregate toxicity extends previous data suggesting that neuronal loss in AD could result from mitotic reactivation of terminally differentiated neurons with arrest in the S phase.

A paradox of cadmium: a carcinogen that impairs the capability of human breast cancer cells to induce angiogenesis.

Cadmium, a highly persistent heavy metal, has been categorized as a human carcinogen. Even though it is known that cadmium acts as estrogens in breast cancer cells, several studies failed to demonstrate whether cadmium is a causal factor for breast cancer. The lack of a strong association between cadmium and breast cancer could be found in the antiangiogenic properties of this heavy metal, which might counteract its carcinogenic properties in the progression of breast cancer. In this study, we exposed estrogen-responsive breast cancer cells to subtoxic levels of cadmium, and we evaluated their angiogenic potential using the chick embryo chorioallantoic membrane assay. Exposure of breast cancer cells to subtoxic levels of cadmium significantly inhibited the angiogenic potential of the breast cancer cell line, suggesting the possibility that cadmium might negatively regulate the production of proangiogenic factors in breast cancer cells. Our results suggest that cadmium might exert a paradoxical effect in breast cancer: on the one hand, it could promote carcinogenesis, and, on the other hand, it could delay the onset of tumors by inhibiting breast cancer cell-induced angiogenesis.

Short-Term Effects of Focal Muscle Vibration on Motor Recovery After Acute Stroke: A Pilot Randomized Sham-Controlled Study.

Repetitive focal muscle vibration (rMV) is known to promote neural plasticity and long-lasting motor recovery in chronic stroke patients. Those structural and functional changes within the motor network underlying motor recovery occur in the very first hours after stroke. Nonetheless, to our knowledge, no rMV-based studies have been carried out in acute stroke patients so far, and the clinical benefit of rMV in this phase of stroke is yet to be determined. The aim of this randomized double-blind sham-controlled study is to investigate the short-term effect of rMV on motor recovery in acute stroke patients. Out of 22 acute stroke patients, 10 were treated with the rMV (vibration group-VG), while 12 underwent the sham treatment (control group-CG). Both treatments were carried out for 3 consecutive days, starting within 72 h of stroke onset; each daily session consisted of three 10-min treatments (for each treated limb), interspersed with a 1-min interval. rMV was delivered using a specific device (Cro®System, NEMOCO srl, Italy). The transducer was applied perpendicular to the target muscle's belly, near its distal tendon insertion, generating a 0.2-0.5 mm peak-to-peak sinusoidal displacement at a frequency of 100 Hz. All participants also underwent a daily standard rehabilitation program. The study protocol underwent local ethics committee approval (ClinicalTrial.gov NCT03697525) and written informed consent was obtained from all of the participants. With regard to the different pre-treatment clinical statuses, VG patients showed significant clinical improvement with respect to CG-treated patients among the NIHSS (p < 0.001), Fugl-Meyer (p = 0.001), and Motricity Index (p < 0.001) scores. In addition, when the upper and lower limb scales scores were compared between the two groups, VG patients were found to have a better clinical improvement at all the clinical end points. This study provides the first evidence that rMV is able to improve the motor outcome in a cohort of acute stroke patients, regardless of the pretreatment clinical status. Being a safe and well-tolerated intervention, which is easy to perform at the bedside, rMV may represent a valid complementary non-pharmacological therapy to promote motor recovery in acute stroke patients.

Heparin/heparan sulfate anticoagulant glycosaminoglycans in human plasma of healthy donors: preliminary study on a small group of recruits.

Glycosaminoglycans in normal human plasma, mainly represented by chondroitin sulfates and heparan sulfates/heparin (HSGAGs), show a specific distribution in the Cohn-Oncley fractions of human plasma. In the present study we investigated their effects on coagulation. Plasma was fractionated following the procedure of Cohn-Oncley, and each fraction was treated for extraction of glycosaminoglycans after extensive proteolysis; the anticoagulant activity in the extracted samples was measured by activated partial thromboplastin time (APTT). The effects of the samples containing HSGAGs on factor II and factor X activities, before and after treatment with heparinase I, were also measured. The molecular weight of HSGAGs was determined by polyacrylamide gel-electrophoresis. Cryoprecipitate and fraction I, fraction II+III, and fraction IV-1 (the fractions containing HSGAGs) prolonged the APTT, whereas fractions IV-4 and V had no effect on the APTT. Fractions containing HSGAGs showed effects on factor II and factor X activities that were sensitive to heparinase I treatment. The molecular weight of HSGAGs recovered in cryoprecipitate and fraction I was 15-18 kDa; that of HSGAGs recovered in fraction IV-1 was 12.0 kDa. In conclusion, these results demonstrate that HSGAGs of different molecular weight, endowed with anticoagulant activity, circulate in normal human plasma in association with specific proteins involved in the regulation of hemostasis; and that endogenous HSGAGs play a role in maintaining the antithrombotic/hemostatic balance in normal human plasma.

Bombesin promotes vasculogenesis and angiogenesis in chick chorio-allantoic membrane: A morphometric, structural, and ultrastructural study.

Experiments were performed on the chorio-allantoic membrane (CAM) of the chick to evaluate the effects of bombesin (BN) on vascular neoformation. In morphometrical assays, 10(-13)-10(-4) M BN promoted dose-dependent vascular development. Newly formed vessels converged toward the BN release site in a spoked wheel arrangement, suggesting a diffusion gradient mechanism. Structural and ultrastructural analysis of CAM specimens collected near the BN release site showed that both vasculogenetic and angiogenetic processes cooperated in vascular neoformation that involved committed cells from the mesenchyme (angioblasts) as well as endothelial cells. No pattern of vascular development was detected away from the BN release site. Findings from the present study emphasize the role of BN in vascular net development of respiratory organs.

Friend erythroleukemia cells induce angiogenesis in chick embryo chorioallantoic membrane and in human umbilical vein endothelial cells.

The effects of Friend erythroleukemia cells on angiogenesis were studied in chick embryo chorioallantoic membrane assay and in human umbilical vein endothelial cells. In chorioallantoic membrane assay, the conditioned medium of Friend cells stimulated in vivo angiogenesis to an extent comparable to that observed with Prostaglandin El, used as positive control. Prostaglandin El added to conditioned medium of Friend cells did not further increase angiogenesis. Conditioned medium of Friend erythroleukemia cells also stimulated proliferation of human umbilical vein endothelial cells to an extent comparable to that observed with fetal bovine serum, used as positive control. Conditioned medium and fetal bovine serum together did not affect human umbilical vein endothelial cells proliferation, as compared to that observed when tested separately. These results seem to indicate that Friend erythroleukemia cells produce and secrete factors stimulating angiogenesis. These findings extend and confirm the hypothesis that successful angiogenesis is necessary for development of leukemias.

Improved method for analysis of glycosaminoglycans in glycosaminoglycan/protein mixtures: application in Cohn-Oncley fractions of human plasma.

BACKGROUND: Glycosaminoglycans are found in human tissues including plasma. They encompass chondroitin sulphates, heparan sulphate/heparin, hyaluronic acid, and keratan sulphate. Glycosaminoglycans, in particular heparan sulphate and heparin, are strongly associated with plasma proteins, so that their purification results quite difficult. METHODS: In order to study the distribution of glycosaminoglycans in plasma subfractions, we developed a novel method that allows their identification even if they were still associated with proteins or peptides. Plasma was fractionated following the procedure of Cohn-Oncley, and each fraction was treated with proteases. After centrifugation, glycosaminoglycan/protein complexes in the supernatant were analysed using a modified cellulose acetate electrophoresis which allowed identification of glycosaminoglycans in mixtures of glycosaminoglycans/proteins. RESULTS: Chondroitin sulphate was recovered in cryoprecipitate and in all Cohn-Oncley fractions. Glycosaminoglycans belonging to the class of heparan sulphate/heparin, however, were recovered in the cryoprecipitate and in fractions I and IV-1, and, in smaller amount, in fraction II+III. CONCLUSIONS: Since the largest amount of plasma proteins is partitioned in Factions II+III and V, these results demonstrate that heparan sulphate/heparin are not randomly distributed in Cohn-Oncley fractions and are associated with certain plasma proteins. This association might play a role in the physiological function of heparan sulphate/heparin, regulating hemostasis and atherogenesis.