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Paolo Madeddu

Publications and source records attributed to Paolo Madeddu.

At least 19 recordsLinked to original sources
PubMed2007

Type-2 diabetic Lepr(db/db) mice show a defective microvascular phenotype under basal conditions and an impaired response to angiogenesis gene therapy in the setting of limb ischemia.

Diabetes mellitus is associated with macro- and micro-angiopathy, leading to increased risk of peripheral ischemia. In the present study, we have characterized the microvascular phenotype at the level of limb muscles and the spontaneous angiogenesis response to surgically-induced unilateral limb ischemia in a murine model of type-2 diabetes, the obese C57BL/KsOlaHsd-Lepr(db/db) mice (Lepr(db/db)), and in non-diabetic heterozygous Lepr(db/+). Wild type C57BL mice (WT) were used as controls. The basal microvascular phenotype was determined in mice aged 3 or 5 months, while the response to limb ischemia was studied only in 5-month old mice. Moreover, in 5-month old ischemic Lepr(db/db) and Lepr(db/+), we have tested the therapeutic potential of local angiogenesis gene therapy with human tissue kallikrein (hTK) or constitutively-activated Akt kinase (Myr-Akt). We found that in the muscles of 3- or 5-month old Lepr(db/db), apoptosis of endothelial cells was enhanced and the densities of capillary and arteriole were reduced. Arterioles of Lepr(db/db) showed hypertrophic remodelling and, occasionally, lumen occlusion. Following ischemia, Lepr(db/db) showed a defective reparative angiogenesis in ischemic muscle, delayed blood flow recovery, and worsened clinical outcome as compared with controls. Five-month old Lepr(db/+) displayed an increase in endothelial cell apoptosis under basal conditions, while capillary and arteriole densities were normal. Lepr(db/+) mounted a proper reparative angiogenesis response to limb ischemia and regained blood flow to the ischemic limb, regularly. Local gene therapy with hTK or Myr-Akt induced angiogenesis in ischemic muscles of Lepr(db/+) and Lepr(db/db). However, in the Lepr(db/db) neither gene therapy approach improved the blood flow recovery and the clinical outcome from ischemia. In contrast, either hTK or Myr-Akt gene transfer improved the post-ischemic recovery of Lepr(db/+). Type-2 diabetes has a negative impact on the basal microvascular phenotype and severely impairs post-ischemic recovery of limb muscles. Gene therapy-induced stimulation of neovascularization might not suffice as a sole therapeutic strategy to combat type-2 diabetes-related vascular complications. In type-2 diabetic patients, therapeutic angiogenesis may need to be further optimized before being recommended for clinical applications.

Animals↗
PubMed2006

Vascular endothelium summary statement VI: Research directions for the 21st century.

Effective translation of research advances from the bench to clinical and public health practice at the bedside and in the community at large represents an important step in the health research discovery enterprise. Increasingly, the gap in translating these advances into practice is being recognized. Successfully addressing this translational gap for the prevention and control of chronic diseases will require the development of novel, innovative, and, if necessary, nontraditional approaches. Participants in the 8th International Conference on Vascular Endothelium discussed a variety of novel approaches that have significant promise. Three of these approaches-vaccine development, genomics and proteomics, and tissue engineering-are highlighted in this position statement and strategies for public health practice and research are suggested.

Animals↗
PubMed2006

Therapeutic angiogenesis: translating experimental concepts to medically relevant goals.

Angiogenesis is central to many physiological and pathological phenomena. In physiological angiogenesis, new vessels are well shaped and their growth is finely tuned to match the metabolic needs of tributary tissues. Accordingly, neovascularization is activated by physical exercise and destabilized by non-use. In contrast, pathological blood vessels that are observed in retinal neovascularization, cancer or in ischemic tissues are leaky, irregularly shaped, and tend to form arterial-venous fistulae. A great deal of attention is focused on new approaches for medical manipulation of vascular growth. These methods are aimed at facilitating the reperfusion of ischemic tissues or eradicating pathological vasculature. In this position paper, we challenge the rationale of therapeutic angiogenesis for the cure of myocardial and peripheral ischemia. Therapeutic angiogenesis aims at combating the insufficiency of, or insensitivity to angiogenic factors in the setting of atherosclerotic-induced arterial occlusion. However, clinical evidence indicates that such a defect is not common among patients with ischemic disease, as a whole. Genetic and environmental factors could account for the great heterogeneity in the expression of the master angiogenic factors. Future improvements in the strategy would require the introduction of in vitro assays and in vivo imaging systems for assessing human angiogenesis. Finally, the promise is to find individualized angiogenesis-based therapies for a genuine cure of ischemia and prevention of organ failure.

Angiogenesis Inducing Agents↗
PubMed2005

Diagnostic tools for the study of vascular cognitive dysfunction in hypertension and antihypertensive drug research.

Arterial hypertension is one of the main risk factors for cerebrovascular diseases, and antihypertensive treatment has significantly reduced their associated mortality. However, morbidity has not been reduced to a similar extent and a still increasing number of patients suffers from recurring strokes and from the disabling consequences of cerebrovascular diseases and develops progressive cognitive impairment. It is still debated to what extent antihypertensive treatment may prevent the development of cognitive dysfunction, due to the lack of a focused approach to vascular cognitive impairment, to the lack of a systematic study of the early phases of dementia, and to the use of diagnostic tests that are not sensitive and specific for a slow onset clinical condition, such as dementia. The aim of the present expert consensus report is to enlist the diagnostic tools that are currently available to assess mild cognitive impairment (MCI) and early dementia and that are sensitive and specific enough to be used in observational, longitudinal, and interventional clinical research studies, aiming to investigate the impact of antihypertensive drugs on vascular dementia (VD).

Animals↗
PubMed2005

Genetic deletion of the p66Shc adaptor protein protects from angiotensin II-induced myocardial damage.

Angiotensin II (Ang II), acting through its G protein-coupled AT1 receptor (AT1), contributes to the precocious heart senescence typical of patients with hypertension, atherosclerosis, and diabetes. AT1 was suggested to transactivate an intracellular signaling controlled by growth factors and their tyrosin-kinase receptors. In cultured vascular smooth muscle cells, this downstream mechanism comprises the p66Shc adaptor protein, previously recognized to play a role in vascular cell senescence and death. The aim of the present study was 2-fold: (1) to characterize the cardiovascular phenotype of p66Shc knockout mice (p66Shc(-/-)), and (2) to test the novel hypothesis that disrupting the p66Shc might protect the heart from the damaging action of elevated Ang II levels. Compared with wild-type littermates (p66Shc(+/+)), p66Shc(-/-) showed similar blood pressure, heart rate, and left ventricular wall thickness. However, cardiomyocyte number was increased in mutant animals, indicating a condition of myocardial hyperplasia. In p66Shc(+/+), infusion of a sub-pressor dose of Ang II (300 nmol/kg body weight [BW] daily for 28 days) caused left ventricular hypertrophy and apoptotic death of cardiomyocytes and endothelial cells. In contrast, p66Shc(-/-) were resistant to the proapoptotic/hypertrophic action of Ang II. Consistently, in vitro experiments showed that Ang II causes apoptotic death of cardiomyocytes isolated from p66Shc(+/+) hearts to a greater extent as compared with p66Shc(-/-) cardiomyocytes. Our results indicate a fundamental role of p66Shc in Ang II-mediated myocardial remodeling. In perspective, p66Shc inhibition may be envisioned as a novel way to prevent the deleterious effects of Ang II on the heart.

Adaptor Proteins, Signal Transducing↗
PubMed2005

Therapeutic angiogenesis and vasculogenesis for tissue regeneration.

Therapeutic angiogenesis/vasculogenesis holds promise for the cure of ischaemic disease. The approach postulates the manipulation of spontaneous healing response by supplementation of growth factors or transplantation of vascular progenitor cells. These supplements are intended to foster the formation of arterial collaterals and promote the regeneration of damaged tissues. Angiogenic factors are generally delivered in the form of recombinant proteins or by gene transfer using viral vectors. In addition, new non-viral methods are gaining importance for their safer profile. The association of growth factors with different biological activity might offer distinct advantages in terms of efficacy, yet combined approaches require further optimization. Alternatively, substances with pleiotropic activity might be considered, by virtue of their ability to target multiple mechanisms. For instance, some angiogenic factors not only stimulate the growth of arterioles and capillaries, but also inhibit vascular destabilization triggered by metabolic and oxidative stress. Transplantation of endothelial progenitor cells was recently proposed for the treatment of peripheral and myocardial ischaemia. Progenitor cells can be transplanted either without any preliminary conditioning or after ex vivo genetic manipulation. Delivery of genetically modified progenitor cells eliminates the drawback of immune response against viral vectors and makes feasible repeating the therapeutic procedure in case of injury recurrence. It is envisioned that these new approaches of regenerative medicine will open unprecedented opportunities for the care of life-threatening diseases.

Animals↗
PubMed2005

Changing the logic of therapeutic angiogenesis for ischemic disease.

The worldwide epidemic of ischemic disease urgently requires innovative treatments. Recently, therapeutic angiogenesis has emerged as a noninvasive supply-side approach, aimed at promoting neovascularization in underperfused tissues through the local delivery of angiogenic growth factors. Successful preclinical studies paved the way for the first clinical trials, with single growth factors given as recombinant proteins or genes. However, clinical results have not matched the initial promises. Our opinion is that the logic of therapeutic angiogenesis needs profound revision. Here, we introduce the concept that pleiotropic agents can stimulate the healing of all the components of ischemic tissue. We also propose prophylactic interventions to delay vascular senescence. The optimization of therapeutic angiogenesis will open unprecedented opportunities for the care of life-threatening ischemic disease.

Angiogenesis Inducing Agents↗
PubMed2005

In search of the best candidate for regeneration of ischemic tissues: are embryonic/fetal stem cells more advantageous than adult counterparts?

Human stem cells and progenitor cells from the bone marrow have been proposed for the regeneration of ischemic cardiac tissues. Early clinical trials indicate that infusion of autologous bone-marrow cells into the infarcted heart enhances ventricular function, albeit the long-term benefit remains to be ascertained. Alternatively, angiogenic growth factors could be used to stimulate the recruitment of vascular progenitor cells into tissues in need of regeneration. Unfortunately, in atherosclerotic patients, the curative potential of autologous stem cells might be impoverished by underlying disease and associated risk factors. Thus, research is focusing on the use of embryonic stem cells which are capable of unlimited self-renewal and have the potential to give rise to all tissue types in the body. Ethical problems and technical hurdles may limit the immediate application of embryonic stem cells. In the meanwhile, fetal hematopoietic stem cells,which have been routinely used to reconstitute the hematopoietic system in man, could represent an alternative, owing to their juvenile phenotype and ability to differentiate into vascular endothelial, muscular, and neuronal cell lineages. With progresses in stem cell expansion, the blood of a single cord could be sufficient to transplant an adult. These observations raise the exciting possibility of using fetal cells as a new way to speed up the healing of damaged tissues.

Cord Blood Stem Cell Transplantation↗
PubMed2004

Akt/protein kinase B and endothelial nitric oxide synthase mediate muscular neovascularization induced by tissue kallikrein gene transfer.

BACKGROUND: Angiogenesis gene therapy with human tissue kallikrein (hTK) has shown promise for ischemic disease. The present study was undertaken to (1) assess an optimal gene transfer modality, (2) clarify hTK angiogenic pathways, and (3) discount possible side effects. METHODS AND RESULTS: The hTK gene was transferred to murine adductors by increasing doses of an adenovirus (Ad.hTK). Heterologous protein production was evaluated by ELISA and immunohistochemistry. Structural and functional characteristics of hTK-induced neovascularization were assessed. Muscular endothelial nitric oxide synthase (eNOS) and vascular endothelial growth factor (VEGF)-A mRNA and protein content were evaluated by real-time polymerase chain reaction and Western blotting. The ability of hTK to phosphorylate-activate Akt/protein kinase B (Akt-B) and VEGF receptor 2 (VEGF-R2) was also determined. Implication of the aforementioned mechanisms in Ad.hTK-induced neovascularization was challenged by blocking Akt-B with a dominant-negative Akt construct; NOS with N(G)-nitro-L-arginine methyl ester; and VEGF-A with neutralizing antibody, VEGF-R2 antagonist, or Ad carrying soluble VEGF-R1 gene. We found that 10(7) PFU Ad.hTK led to peak increases in capillary and arteriole density. Newly developed arterioles persisted for up to 8 weeks. Ad.hTK did not change microvascular permeability. Ad.hTK upregulated eNOS mRNA and protein and activated Akt-B through Ser-473 phosphorylation. Inhibitory studies documented that these biochemical events were instrumental to Ad.hTK-induced neovascularization. In contrast, Ad.hTK neither affected VEGF-A and VEGF-R2 levels nor increased VEGF-R2 phosphorylation. Consistently, Ad.hTK-induced neovascularization was not disturbed by any of the different approaches used to block VEGF-A. CONCLUSIONS: Our findings provide new information on the pathway involved in hTK-induced neoangiogenesis and represent an advancement toward clinical applications with Ad.hTK.

Adenoviridae↗
PubMed2004

Transplantation of low dose CD34+KDR+ cells promotes vascular and muscular regeneration in ischemic limbs.

Hematopoietic progenitor cell transplantation can contribute to revascularization of ischemic tissues. Yet, the optimal cell population to be transplanted has yet to be determined. We have compared the therapeutic potential of two subsets of human cord blood CD34+ progenitors, either expressing the VEGF-A receptor 2 (KDR) or not. In serum-free starvation culture, CD34+KDR+ cells reportedly showed greater resistance to apoptosis and ability to release VEGF-A, as compared with CD34+KDR- cells. When injected into the hind muscles in immunodeficient SCIDbg mice subjected to unilateral ischemia, a low number (10(3)) of CD34+KDR+ cells improved limb salvage and hemodynamic recovery better than a larger dosage (10(4)) of CD34+KDR- cells. The neovascularization induced by KDR+ cells was significantly superior to that promoted by KDR- cells. Similarly, endothelial cell apoptosis and interstitial fibrosis were significantly attenuated by KDR+ cells, which differentiated into mature human endothelial cells and also apparently skeletal muscle cells. This study demonstrates that a low number of CD34+KDR+ cells favors reparative neovascularization and possibly myogenesis in limb ischemia, suggesting the potential use of this cell population in regenerative medicine.

Animals↗
PubMed2004

Nitric oxide-releasing aspirin derivative, NCX 4016, promotes reparative angiogenesis and prevents apoptosis and oxidative stress in a mouse model of peripheral ischemia.

BACKGROUND: Recently, nitric oxide (NO) donors have been developed that mimic the physiological intracellular release of NO. We evaluated whether one of these new compounds, consisting of aspirin coupled to an NO-releasing moiety (NCX 4016), would protect limbs from supervening arterial occlusion. METHODS AND RESULTS: Mice were assigned to receive regular chow or chow containing NCX 4016 or aspirin (both at 300 mumol/kg body weight, daily) throughout the 3-week experimental period. One week after randomization, they underwent surgical excision of the left femoral artery. Limb blood flow recovery (laser Doppler flowmetry) was accelerated by NCX 4016 as compared with aspirin or vehicle (P<0.05). In controls, histological analysis revealed a 35% increase in the capillary density of ischemic muscles compared with contralateral ones, indicative of spontaneous angiogenesis. Neovascularization was enhanced by NCX 4016 (91%; P<0.05 versus vehicle), but not by aspirin (51%; P=NS versus vehicle). Furthermore, NCX 4016 reduced endothelial cell (EC) apoptosis (4.3+/-1.0 versus 8.7+/-2.0 in aspirin and 12.6+/-3.3 ECs/1000 cap in vehicle; P<0.05 for either comparison) as well as caspase-3 mRNA levels in ischemic muscles ([caspase-3/GAPDH]*100 = 0.09+/-0.04 versus 2.30+/-0.44 in aspirin and 2.30+/-0.32 in vehicle; P<0.01 for either comparison). Nitrite levels and the ratio of reduced to oxidized glutathione were selectively increased in ischemic muscles by NCX 4016. Vascular endothelial growth factor-A expression was reduced by aspirin, with this effect being blunted by NCX 4016. CONCLUSIONS: Pretreatment with the new oral NO-releasing aspirin derivative stimulates reparative angiogenesis and prevents apoptosis and oxidative stress, thereby alleviating the consequences of supervening arterial occlusion.

Animals↗
PubMed2004

Circulating tissue kallikrein levels correlate with severity of carotid atherosclerosis.

BACKGROUND: Vascular growth factors are upregulated in stroke patients, but it remains unknown if they correlate with carotid atherosclerosis. METHODS AND RESULTS: A case-control study was conducted to determine: (1) possible association between biomarkers of angiogenesis or inflammation and carotid stenosis; and (2) the impact of revascularization on the same biomarkers. Circulating vascular endothelial growth factor (VEGF), basic fibroblast GF (bFGF), tissue kallikrein (tK), and high-sensitivity C-reactive protein (hs-CRP) were measured in 89 patients with carotid obstruction and 45 age-matched controls. Patients were stratified as <50% carotid stenosis (CAS; n=16); 50% to 69% CAS (n=12); 70% to 99% CAS (n=43); and carotid occlusion (CAO; n=18). No association was found between VEGF, bFGF, or hs-CRP and obstruction grading. TK augmented from 360+/-30 in <50% CAS (P=NS versus controls) to 509+/-72 in moderate CAS (P<0.05), 1159+/-178 in high-grade CAS (P<0.02), and 1616+/-403 pg/mL in CAO (P<0.01). A threshold of 508 pg/mL provided the maximized predictive value of high-grade obstruction. After revascularization, tK decreased from 1410+/-352 to 782+/-86 pg/mL (P<0.01), whereas no change was detected in nonoperated cases. Hs-CRP was unaffected by revascularization. CONCLUSIONS: Angiogenic factors are heterogeneously expressed in patients with carotid atherosclerosis. The tK measurement may be useful for the diagnosis and monitoring of atherosclerotic disease.

Adult↗
PubMed2004

Genetically engineered stem cell therapy for tissue regeneration.

Therapeutic angiogenesis/vasculogenesis represents a new approach to treat patients with ischemic disease not curable with conventional treatment. This review focuses on the rationale and preliminary results of combining stem cell and gene therapy for regenerative medicine. Under disease conditions, impaired neovascularization results from diminished vascular growth factor production and primary dysfunction of endothelial cells and their progenitors. Advances in our ability to genetically manipulate cells ex vivo has provided the technological platform to implement stem cell biology and circumvent the potential hazard of direct gene transfer. Ex vivo engineered endothelial progenitor cells have been used for the treatment of peripheral limb ischemia. The approach eliminates the drawback of immune response against viral vectors and makes feasible repeating the therapeutic procedure in case of injury recurrence. The strategy of using stem cells as vectors for curative agents proved to be of value for the treatment of pulmonary hypertension and thrombosis. Transplantation of neural stem cells genetically modified to secrete nerve growth factor was able to ameliorate the death of striatal projection neurons caused by transient focal ischemia in the adult rat. By a similar approach, engineered neural stem cells might be used for treating neurodegenerative disorders. Therefore, genetic manipulation of stem cells opens new avenues for regenerative medicine.

Animals↗
PubMed2004

Prophylactic gene therapy with human tissue kallikrein ameliorates limb ischemia recovery in type 1 diabetic mice.

Diabetes macro- and microvascular disease causes tissue hypoperfusion. This deficit, together with a failure to mount an adequate angiogenic response, might explain why vascular occlusion evolves more severely among diabetic patients. The present study investigated whether prophylactic gene therapy with human tissue kallikrein (hTK) may protect diabetic limbs from the consequences of supervening ischemia. Vehicle (saline) or an adenovirus carrying the gene for either hTK (Ad.hTK) or luciferase (Ad.Luc) was injected into left adductor muscles of streptozotocin-induced type 1 diabetic mice 2 weeks before operative occlusion of the ipsilateral femoral artery. Saline-injected nondiabetic mice served as controls. Hindlimb blood flow recovery was analyzed sequentially over the 2 weeks after ischemia induction. At necroscopy, microvessel density and endothelial cell proliferation and apoptosis were quantified in skeletal muscles. We found that limb perfusion recovery of saline-injected type 1 diabetic mice is delayed because of insufficient reparative neovascularization and excessive activation of endothelial cell apoptosis. By contrast, prophylactic Ad.hTK renewed the ability to mount an appropriate neovascularization response to ischemia, suppressed apoptosis, and upregulated endothelial nitric oxide synthase expression. Ultimately, correction of diabetic endotheliopathy by Ad.hTK allowed proper perfusion recovery as seen in nondiabetic mice. These discoveries disclose new therapeutic options for the treatment of diabetic complications.

Animals↗
PubMed2003

Paracrine control of vascularization and neurogenesis by neurotrophins.

The neuronal system plays a fundamental role in the maturation of primitive embryonic vascular network by providing a paracrine template for blood vessel branching and arterial differentiation. Furthermore, postnatal vascular and neural regeneration cooperate in the healing of damaged tissue. Neurogenesis continues in adulthood although confined to specific brain regions. Following ischaemic insult, neural staminal cells contribute towards the healing process through the stimulation of neurogenesis and vasculogenesis. Evidence indicates that nerves and blood vessels exert a reciprocal control of their own growth by paracrine mechanisms. For instance, guidance factors, including vascular endothelial growth factor A (VEGF-A) and semaphorins, which share the ability of binding neuropilin receptors, play a pivotal role in the tridimensional growth pattern of arterial vessels and nerves. Animal models and clinical studies have demonstrated a role of VEGF-A in the pathogenesis of ischaemic and diabetic neuropathies. Further, supplementation with VEGF-A ameliorates neuronal recovery by exerting protective effects on nerves and stimulating reparative neovascularization. Human tissue kallikrein, a recently discovered angiogenic and arteriogenic factor, accelerates neuronal recovery by stimulating the growth of vasa nervorum. Conversely, the neurotrophin nerve growth factor, known to regulate neuronal survival and differentiation, is now regarded as a stimulator of angiogenesis and arteriogenesis. These results indicate that angiogenesis and neurogenesis are paracrinally regulated by growth factors released by endothelial cells and neurons. Supplementation of these growth factors, alone or in combination, could benefit the treatment of ischaemic diseases and neuropathies.

Humans↗
PubMed2003

Cardiac hypertrophy and microvascular deficit in kinin B2 receptor knockout mice.

Experimental and clinical evidence suggests kinin involvement in adaptive myocardial growth. Kinins are growth-inhibitory to cardiomyocytes. Knockout of kinin B2 receptor (B2R) signaling causes dilated and failing cardiomyopathy in 129/J mice, and a 9-bp deletion polymorphism of human B2R is associated with reduced receptor expression and exaggerated left ventricular growth response to physical stress. We reasoned that genetic background and aging may significantly influence the impact of B2R mutation on cardiac phenotype. The theory was challenged in C57BL/6 mice, a strain that naturally differs from the 129/J strain, carrying 1 instead of 2 renin genes. C57BL/6 B2R knockouts (B2R-KO) showed higher blood pressure and heart rate levels (P<0.05) compared with wild-type controls (WT) at all ages examined. At 12 months, left ventricular contractility and diastolic function were mildly altered (P<0.05) and histological and morphological analyses revealed ventricular hypertrophy and cardiomyocyte enlargement in B2R-KO (P<0.01). Reparative fibrosis was enhanced by 208% and capillary density reduced by 38% (P<0.01). Functional and structural alterations induced by B2R deletion in C57BL/6 mice were less severe than those reported previously in the 129/J strain. We conclude that interaction of B2R signaling with other genetic determinants influences aging-related changes in myocardial structure and function. These findings may help us understand the role of kinins in the development of cardiac failure.

Animals↗
PubMed2003

Human tissue kallikrein: a new bullet for the treatment of ischemia.

Recently, therapeutic angiogenesis has been proposed as an alternative for the treatment of ischemic diseases unresponsive to conventional therapy. This strategy is based on the concept that a supply-side approach with growth factors would overcome the endogenous deficit and result in more robust collateralization. We have developed a strategy based on local delivery of human tissue kallikrein gene for potentiation of microcirculation and rescue of peripheral ischemia. Following successful application in otherwise healthy animals, the approach resulted to be of therapeutic value in rats with endothelial dysfunction caused by arterial hypertension. In addition, human tissue kallikrein prevents or rescues microvascular rarefaction caused by diabetes mellitus. In this model, human tissue kallikrein was able to stimulate vascular growth and contrast apoptosis. The strategy displays interesting pharmacological features because is devoid of obvious side effects and is effective even at low infecting doses. In addition, the neovascularization promoted by human tissue kallikrein is well organized and durable. It is reasonable to anticipate that the new approach will have a great impact in the treatment of cardiovascular ischemic complications.

Animals↗