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ABSTRACT: We studied the gene therapy efficacy of diabetes-associated wound healing disorder with an adeno-associated virus (AAV) vector expressing the 165-amino acid isoform of human vascular endothelial growth factor-A (VEGF-A) by using an incisional skin-wound model produced on the back of female diabetic C57BL/KsJ db+/db+ mice and their normal littermates ( db+/+m).
Animals were randomized to receive intradermally into the wound edges either rAAV-LacZ (a control gene), or rAAV-VEGF165. Animals were killed on different days (7 and 14 days after skin injury) and wounded skin tissues were used for gene marker studies, histological evaluation and immunohistochemistry, and wound breaking strength analysis. Furthermore we studied the VEGF mature protein in the wounds.
We found that AAV vectors are highly efficient for gene transfer to the mouse skin, displaying an exquisite tropism for the panniculus carnosus by using the beta-galactosidase activity assay. We confirmed the increased expression of the angiogenic factor at day 7 by measuring the wound content of the mature protein. Delivery of VEGF165 to incisional skin wounds of diabetic mice resulted in a remarkable induction of new vessel formation with consequent improvement in the wound healing process. The rAAV-VEGF165 gene improved wound healing in diabetic mice through the stimulation of angiogenesis, reepithelization, synthesis and maturation of extracellular matrix. Moreover the recombinant AAV encoding the human VEGF165 increased the breaking strength of the wound and enhanced the wound content of VEGF.
Our study suggests that VEGF gene transfer might represent a new approach to treat wound healing disorders associated with diabetes.
Diabetologia 05/2003; 46(4):546-55. · 6.81 Impact Factor
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B Deodato,
N Arsic,
L Zentilin,
M Galeano,
D Santoro, V Torre,
D Altavilla,
D Valdembri,
F Bussolino,
F Squadrito,
M Giacca
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ABSTRACT: Delivery of therapeutic genes represents an appealing possibility to accelerate healing of wounds that are otherwise difficult to treat, such as those in patients with metabolic disorders or infections. Experimental evidence indicates that in such conditions potentiation of neo-angiogenesis at the wound site might represent an important therapeutic target. Here we explore the efficacy of gene therapy of wound healing with an adeno-associated virus (AAV) vector expressing the 165 amino acid isoform of vascular endothelial growth factor-A (VEGF-A). By gene marker studies, we found that AAV vectors are highly efficient for gene transfer to the rat skin, displaying an exquisite tropism for the panniculus carnosus. Gene expression from these vectors is sustained and persistent over time. Delivery of VEGF165 to full thickness excisional wounds in rats resulted in remarkable induction of new vessel formation, with consequent reduction of the healing time. Histological examination of treated wounds revealed accelerated remodeling of epidermis and dermis, with formation of a thick granular layer, containing numerous newly formed capillaries, as well as vessels of larger size. These data underline the importance of neo-angiogenesis in the healing process and indicate that VEGF gene transfer might represent a novel approach to treat wound healing disorders.
Gene Therapy 07/2002; 9(12):777-85. · 3.71 Impact Factor
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M Galeano, V Torre,
B Deodato,
G M Campo,
M Colonna,
A Sturiale,
F Squadrito,
V Cavallari,
D Cucinotta,
M Buemi,
D Altavilla
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ABSTRACT: Impaired wound healing is a well-documented phenomenon in experimental and clinical diabetes. Emerging evidence favors the involvement of free radicals in the pathogenesis of diabetes-related healing deficit. This study assessed the effect of systemic administration of raxofelast, a protective membrane antioxidant agent, on wound healing by using healing-impaired (db/db) mice.
The wound healing effect of raxofelast was investigated by using an incisional skin-wound model produced on the back of female diabetic C57BL/KsJ db+/db+ mice and their healthy littermates (db+/+m). Animals were then randomized to the following treatment: raxofelast (15 mg/kg/d intraperitoneally) or its vehicle (dimethyl sulfoxide/sodium chloride 0.9%, 1:1, vol/vol). The animals were killed on different days, and the wounded skin tissues were used for histologic evaluation and for analysis of malondialdehyde (MDA) level and myeloperoxidase (MPO) activity, wound breaking strength, and collagen content.
Diabetic mice showed delayed wound healing together with low collagen content, breaking strength, and increased MDA levels and MPO activity when compared with their healthy littermates. The administration of raxofelast did not modify the process of wound repair in healthy (db/+) mice, but significantly improved impaired wound healing in diabetic mice through the stimulation of angiogenesis, reepithelialization, synthesis, and maturation of extracellular matrix. Furthermore, raxofelast treatment significantly reduced MDA levels, MPO activity, and increased the breaking strength and collagen content of the wound.
The current study provides evidence that raxofelast restores wound healing to nearly normal levels in experimental diabetes-impaired wounds and suggests that an increased lipid peroxidation in diabetic mice may have a role in determining a defect of wound repair.
Surgery 05/2001; 129(4):467-77. · 3.10 Impact Factor
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D Altavilla,
A Saitta,
D Cucinotta,
M Galeano,
B Deodato,
M Colonna, V Torre,
G Russo,
A Sardella,
G Urna,
G M Campo,
V Cavallari,
G Squadrito,
F Squadrito
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ABSTRACT: Impaired wound healing is a well-documented phenomenon in experimental and clinical diabetes. Experimental evidence suggests that a defect in vascular endothelial growth factor (VEGF) regulation might be associated with wound-healing disorders. We studied the involvement of lipid peroxidation in the pathogenesis of altered VEGF expression in diabetes-related healing deficit by using an incisional skin-wound model produced on the back of female diabetic C57BL/KsJ db+/ db+ mice and their normal (db+/+m) littermates. Animals were then randomized to the following treatment: raxofelast (15 mg.kg(-1).day(-1) i.p.), an inhibitor of lipid peroxidation, or its vehicle (DMSO/NaCl 0.9%, 1:1 vol: vol). The animals were killed on different days (3, 6, and 12 days after skin injury), and the wounded skin tissues were used for histological evaluation, for analysis of conjugated dienes (CDs), as an index of lipid peroxidation and wound breaking strength. Furthermore, we studied the time course of VEGF mRNA expression throughout the skin-repair process (3, 6, and 12 days after skin injury), by means of reverse transcriptase-polymerase chain reaction, as well as the mature protein in the wounds. Diabetic mice showed impaired wound healing with delayed angiogenesis, low breaking strength, and increased wound CD content when compared with their normal littermates. In healthy control mice, a strong induction of VEGF mRNA was found between day 3 and day 6 after injury, while no significant VEGF mRNA expression was observed at day 12 after injury. In contrast, VEGF mRNA levels, after an initial increase (day 3), were significantly lower in diabetic mice than in normal littermates, and light induction of VEGF mRNA expression was also present at day 12 after injury. Similarly, the wound content of the angiogenic factor was markedly changed in diabetic mice. Administration of raxofelast did not modify the process of wound repair in normal mice, but significantly improved the impaired wound healing in diabetic mice through the stimulation of angiogenesis, re-epithelization, and synthesis and maturation of extracellular matrix. Moreover, raxofelast treatment significantly reduced wound CD levels and increased the breaking strength of the wound. Lastly, the inhibition of lipid peroxidation restored the defect in VEGF expression during the process of skin repair in diabetic mice and normalized the VEGF wound content. The current study provides evidence that lipid peroxidation inhibition restores wound healing to nearly normal levels in experimental diabetes-impaired wounds and normalizes the defect in VEGF regulation associated with diabetes-induced skin-repair disorders.
Diabetes 04/2001; 50(3):667-74. · 8.29 Impact Factor
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ABSTRACT: We report a case of idiopathic midline destructive disease in a 57-year-old man. The patient had a non-specific histological pattern in biopsies obtained from the nose and upper lip, characterized by a granulomatous reaction with progressive destruction of the tissues. The patient's general medical history was non-contributory. Clinical and laboratory data did not support any feasible etiology for this destructive process. The patient was treated with prednisone until the discovery of type II diabetes mellitus (never diagnosed before) and was then in turn treated only with oral antidiabetic therapy. Follow-up controls revealed progressive reduction of the symptoms and of the nasal and lip lesions and total remission of symptoms up to 2 years after the onset of the disease. We discuss the diagnostic and subsequent therapeutic problems in the management of the midline necrotizing lesions.
Journal of Oral Pathology and Medicine 04/2001; 30(3):190-2. · 1.63 Impact Factor
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ABSTRACT: Primary adenocarcinoma of the larynx is a rare neoplasm that tends to spread to both regional lymph nodes and distant sites. A case of primary adenocarcinoma of the arytenoid in a 74-year-old man is presented. The tumor was evaluated by light and electron microscopy. A high percentage of intranuclear pseudoinclusions (more than 20% of the neoplastic cells) was a peculiar characteristic of the tumor. To the best of our knowledge, such a feature has not been reported previously and should be considered a hallmark of more aggressive behavior.
Pathology - Research and Practice 02/2001; 197(6):449-52. · 1.21 Impact Factor
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ABSTRACT: Eruptive syringomas are uncommon in the general population. We describe here an 18-year-old female, affected by Down's syndrome, who presented with an abrupt eruption of small skin-colored or reddish papules on the face, neck and limbs. Light microscopy allowed us to diagnose syringomas, whereas the study of the ultrastructural features revealed calcium deposits in many lumina and also in the mitochondria. This observation confirms the hypothesis that the syringeal structure plays a role in the pathogenesis of calcinosis cutis.
Dermatology 02/2001; 203(4):345-7. · 2.05 Impact Factor
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ABSTRACT: For poorly differentiated rhinopharyngeal carcinomas, the clinical presentation (association with the Epstein-Barr virus, paraneoplastic syndromes, onset of lymphoma) and the histopathological features can be polymorphous and they can confound or delay diagnosis and preparation of an adequate treatment plan (radio-chemotherapy). Often these neoplasms arise as clinically primitive laterocervical metastases, masked by clinical findings and a history that can lead to the mistaken diagnosis of systemic lymphoproliferative processes such as Hodgkin's disease. Here an observation of this type is presented in a young patient (19 years old) who came under observation for a laterocervical tumefaction recurrent from a previous exeresis performed at another hospital and symptoms of serotine febricula, dysphagia and serology positive for the Epstein-Barr virus (EBV). The patient underwent surgery and then radiotherapy and has been under close post-operative follow-up for two years. To date the patient's condition--both local and general--is good. The particular histology of the neoformation lies in the abundant infiltration of plasma cell and lymphocyte eosinophils, at times in blastic form. Moreover, elements with a large clear nucleus and evident nucleolus (Hodgkin-like) and scattered multinucleate Langhans-type giant cells can be seen. Immunohistologically the tumor cells markedly express for cytokeratin and the latent membrane protein (LMP1) of the Epstein-Barr virus (EBV) and show a high growth fraction. Under the electron microscope, the plurinucleate giant cells present large nuclei with morphology similar to that of tumor cells. The clear cytokeratin-positivity of the tumor elements and the histological and ultrastructural features mentioned led to the diagnosis of a massive metastasis from lymphoepithelial carcinoma, the Schmincke variant, plus EBV infection of the neoplastic cells. The authors conclude assuming that the particular granulomatous reaction is due to the host's reaction to the tumor cells, but also to the reaction to the viral antigens. In the former case we find an attempt to limit the carcinomatous process; in the latter it is a response caused by the EBV and is not, apparently, aimed at protecting against the neoplasm rather it facilitates the neoplastic process.
Acta otorhinolaryngologica Italica: organo ufficiale della Società italiana di otorinolaringologia e chirurgia cervico-facciale 11/2000; 20(5):347-53. · 0.86 Impact Factor
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ABSTRACT: Synovial sarcomas account for 7-10% of all soft tissue malignancies and the rare head and neck region location accounts for an average 5% of them. A brief review of the data in the Literature has shown that not more than one hundred cases of cervical-facial synovial sarcoma have been reported. In the head and neck locations this form of tumor is significantly less aggressive, with a higher survival rate and a recurrence rate much lower than the 60-70% shown for other locations in the limbs. After a brief review of the Literature, Authors present a rare clinical case of hypopharyngeal synovial sarcoma which recently came under observation. Despite the accurate diagnostic procedure, after surgical removal of the tumor by CO2 laser microlaryngoscopy, diagnosis required the use of histological and ultrastructural techniques. CT and NMR of the neck using contrast medium showed that the structure of the neoformation was similar to that of the soft tissues, it was 3-4 cm in diameter, located in the hypopharynx and had an extensive surface in contact with the left glossoepiglottic groove. The lesion appeared to involve the free edge of the aryepiglottic fold, coming into contact with the free edge of the epiglottis; it could not be dissociated form the epiglottis and obliterated both the homolateral glossoepiglottic vallecula and the pyriform sinus. Structural analysis after radiography with a contrast medium showed a marked impregnation of the lesion, indicative of high degree of vascolarization. Ultrastructural, cytofluormetric and immunohistochemical analyses were performed on the neoplasm in order to process all these data together with the clinical parameters; in other words a multi-parameter evaluation was performed, as suggested by other Authors, to determine the therapy and arrange more accurate monitoring of this patient, victim of a neoplasm with a high potential for metastases.
Acta otorhinolaryngologica Italica: organo ufficiale della Società italiana di otorinolaringologia e chirurgia cervico-facciale 09/2000; 20(4):273-80. · 0.86 Impact Factor
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ABSTRACT: According to the "Global Tuberculosis Control" performed in 1999--the third complete, international, global report on tuberculosis infection--173 countries reported their infection data to the WHO; of these countries 102 met the criteria for "DOTS programs" at the end of 1997. The DOTS programs are the only control strategy able to produce a cure rate of 85%. Both at the national and international (Centers of Disease Control) levels, guidelines have been drawn up to improve and coordinate the fight against tuberculosis. New indicators and methods of analysis should be developed to quantify the full impact on the control of infection transmission, incidence, prevalence, mortality and prevention of drug resistance. In addition, two significant world-wide events have affected the increase morbidity rate seen in the last decade in the more highly industrial countries: immigration from countries outside the European Community and HIV infection. The tuberculosis infection worsens the evolution of HIV, facilitating viral replication. In the present work the authors discuss the most recent epidemiological data regarding tuberculosis infection and review the Literature on the primary laryngeal location of the disease. Then they present a clinical case which recently came under observation. This case is a typical example of the clinical picture of the laryngeal tuberculosis seen today. It must not be forgotten that in recent years there has been an increase in morbidity in Italy, in both the pulmonary and extrapulmonary forms of the disease, although in our country the problem of delayed or incomplete reporting is quite widespread. The data show that the age ranges with the highest incidence of both pulmonary and extrapulmonary forms are the 25-35 and 60-70 year groups. Distribution by sex, on the hand, shows that the pulmonary forms are most often seen in males while the extrapulmonary forms have practical the same frequency in both sexes. In recent years the clinical and morphological aspects of tubercular laryngitis have changed significantly from what they were before chemotherapy and the most common clinical form is pseudotumoral tuberculosis. This form requires a differential diagnosis to distinguish it from neoplasms because they present a similar objective picture and have no signs of simultaneous or previous pulmonary involvement.
Acta otorhinolaryngologica Italica: organo ufficiale della Società italiana di otorinolaringologia e chirurgia cervico-facciale 07/2000; 20(3):196-201. · 0.86 Impact Factor
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Pathologica 05/2000; 92(2):129-30.
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Pathologica 05/2000; 92(2):134.
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Pathologica 05/2000; 92(2):134-5.
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ABSTRACT: This study examined 71 pediatric tonsillectomy patients through accurate case history and clinical examination, placing particular emphasis on pathologies concomitant to tonsillopathy. In an attempt to find anatomo-clinical correlations, these data were processed together with the results of a histomorphological study of thetonsil epithelium, performed on all tonsillectomy samples. The majority of these patients were females and none more than 13 years of age. Numerous pathologies were found associated with the tonsillopathy and in varying combinations, first and foremost of which was adenoid hypertrophy. Only approximately one fifth of the patients did not show any concomitant pathology of note. All patients presented a history of recurrent pharyngotonsillitis (at least 4 episodes a year) with symptoms arising from 1 to 10 years prior to surgery. The concomitant pathologies included: respiratory, cutaneous and food allergies, asthma, obstructive sleep apnea, rheumatic diseases, etc. From the histomorphological point of view, particular modifications were found in the follicle epithelium and interstitial cells of the palatine tonsil. An exasperated fibrotic interstitial reaction and chronic duration of the disease appeared to prevent tonsil filter function, facilitating chronicization of the tonsillopathy or onset of recurrent infections and concomitant allergies. In the allergic patients the tonsil epithelium was thickened and compact and showed various degrees of chorion edema, in agreement with what is found in the literature. On the contrary, few morphostructural palatine tonsil mutations were found in those subjects which did not present any concomitant pathology or were affected by tonsillopathy of brief duration. All the histomorphological modifications encountered appear related to the individual patient history, confirming the hypothesis that tonsil epithelium can not only condition the evolution of tonsillopathy--reflecting the effect of various factors--but, above all, it directs the immune response, thus playing a role in the development of various concomitant pathologies.
Acta otorhinolaryngologica Italica: organo ufficiale della Società italiana di otorinolaringologia e chirurgia cervico-facciale 03/2000; 20(1):40-6. · 0.86 Impact Factor
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ABSTRACT: Schwannoma is a rare neoplasm in the E.N.T. areas although it is characteristic of this discipline. Indeed, examination of the most recent data in the literature shows that approximately 45% of all neoplasms of the peripheral nerve linings occur in the head and neck district, the most common neurogenic tumors are schwannomas and these constitute 35% of all head and neck tumors. In reviewing the literature attention was focused on schwannomas originating in unusual sites, neglecting the most common sites originating in the eighth and seventh pair of cranial nerves. It was seen that the site of origin and clinical manifestation of these lesions varies widely and the four cases presented here are no exception. These are four schwannomas which came under observation between 1994 and 1997. The sites of origin were, respectively: the floor of the mouth, the submandibular region (corresponding to the Warthon duct), the anterior wall of the external auditory canal and the hypopharynx (corresponding to the pyriform sinus). All of these cases proved quite rare in the literature. All the cases underwent accurate anatomopathological examination and the paper discusses the particular histological and immunohistochemical features encountered. Reference is also made to the problems of differential diagnosis vs. other types of soft tissue tumors. Emphasis is placed on the demonstrated difficulty in recognizing schwannomas from the macroscopic and surgical points of view. The absence of mitoses, necroses, invasiveness and specific features--i.e. hyperchromia and pleomorphism of the nuclei or the presence of large atypical cells--are all parameters confirming that the lesions observed in the present study were benign. No anatompathological features were observed that could justify any particular expression of the schwannoma in the specific sites involved. Nevertheless, the authors present these case because a review of the literature indicated that they are extremely rare and because differential diagnosis of these unusual manifestations is so complex.
Acta otorhinolaryngologica Italica: organo ufficiale della Società italiana di otorinolaringologia e chirurgia cervico-facciale 07/1999; 19(3):160-5. · 0.86 Impact Factor
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ABSTRACT: Seven cases of primary macular amyloidosis were studied on skin biopsies. The Congo red stain was positive only in three cases, whereas the ultrastructural observation allowed for the detection of amyloid deposits in all biopsies. Fibrillary degeneration of basal keratynocytes was occasionally observed, and regressive changes of keratynocytes and dermal nerve bundles presumably related to the intensity of the scratch trauma were detected in one case. In six biopsies mast cell profiles exhibiting various degrees of degranulation were detected in the dermis. Melanosome aggregates were also observed consistently in dermal macrophages and occasionally in Schwann cells. A variable degree of structural alteration was observed in dermal unmyelinated nerve fibers. Even if the intimate mechanism of amyloid deposition was not explained by the ultrastructural study, this approach is a useful instrument in the differential diagnosis of cutaneous macular hyperpigmented lesions.
Ultrastructural Pathology 23(5):279-84. · 0.76 Impact Factor
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ABSTRACT: Erythema ab igne (EI) is an uncommon skin lesion caused by mild and repeated exposure to infrared sources. The aim of this study was to investigate the ultrastructural alterations in this condition. The ultrastructural study was carried out on 5-outpatients who presented typical EI of their exposed sites. Skin punch biopsies were processed for standard electron microscopy. The epidermis was hyperpigmented, with focal regressive changes of basal keratinocytes. An apparent functional activation of melanocytes with numerical increase of dendritic processes was also observed. The dermis showed abundant melanophages and occasional elastic fiber alterations similar to actinic elastosis. No alterations consistent with preneoplastic skin conditions were observed. The ultrastructural findings associated with EI seem to be nonspecific and consistent with moderate regressive changes of keratinocytes as well as a consensual melanocytic activation and elastic fiber modifications. Similar alterations can be observed in chronic actinic skin damage. This condition is presumably more benign than the ultraviolet exposure. The association of EI and premalignant skin lesions, though occasionally described, seems relatively infrequent.
Ultrastructural Pathology 25(2):93-7. · 0.76 Impact Factor
