Biomedical subjects

P Madeddu

Publications and source records attributed to P Madeddu.

At least 19 recordsLinked to original sources

PubMed2007

Microfabrication of fractal polymeric structures for capillary morphogenesis: applications in therapeutic angiogenesis and in the engineering of vascularized tissue.

Microfabrication techniques were combined with fractal algorithms to realize polymeric scaffolds resembling capillary networks. The scaffolds were seeded with human endothelial cells in monoculture as well as in coculture with human fibroblasts. To enhance the process of angiogenesis, endothelial cells were transfected with an adenoviral vector carrying the gene for human tissue kallikrien. The results demonstrate that both the presence of a structured scaffold as well as fibroblasts in coculture contribute synergically to the promotion of a metabolically active network. The fractal scaffolds have several possible applications for example in vascularized tissue engineering and therapeutic angiogenesis. A broader implication of these results is that cell-extra cellular matrix and cell-cell interactions cooperate dynamically both at a biochemical as well as microstructural level.

Algorithms↗

PubMed2006

Murine models of myocardial and limb ischemia: diagnostic end-points and relevance to clinical problems.

Ischemic disease represents the new epidemic worldwide. Animal models of ischemic disease are useful because they can help us to understand the underlying pathogenetic mechanisms and develop new therapies. The present review article summarizes the results of a consensus conference on the status and future development of experimentation in the field of cardiovascular medicine using murine models of peripheral and myocardial ischemia. The starting point was to recognize the limits of the approach, which mainly derive from species- and disease-related differences in cardiovascular physiology. For instance, the mouse heart beats at a rate 10 times faster than the human heart. Furthermore, healing processes are more rapid in animals, as they rely on mechanisms that may have lost relevance in man. The main objective of the authors was to propose general guidelines, diagnostic end points and relevance to clinical problems.

Animal Experimentation↗

PubMed2006

Benfotiamine accelerates the healing of ischaemic diabetic limbs in mice through protein kinase B/Akt-mediated potentiation of angiogenesis and inhibition of apoptosis.

AIMS/HYPOTHESIS: Benfotiamine, a vitamin B1 analogue, reportedly prevents diabetic microangiopathy. The aim of this study was to evaluate whether benfotiamine is of benefit in reparative neovascularisation using a type I diabetes model of hindlimb ischaemia. We also investigated the involvement of protein kinase B (PKB)/Akt in the therapeutic effects of benfotiamine. METHODS: Streptozotocin-induced diabetic mice, given oral benfotiamine or vehicle, were subjected to unilateral limb ischaemia. Reparative neovascularisation was analysed by histology. The expression of Nos3 and Casp3 was evaluated by real-time PCR, and the activation state of PKB/Akt was assessed by western blot analysis and immunohistochemistry. The functional importance of PKB/Akt in benfotiamine-induced effects was investigated using a dominant-negative construct. RESULTS: Diabetic muscles showed reduced transketolase activity, which was corrected by benfotiamine. Importantly, benfotiamine prevented ischaemia-induced toe necrosis, improved hindlimb perfusion and oxygenation, and restored endothelium-dependent vasodilation. Histological studies revealed the improvement of reparative neovascularisation and the inhibition of endothelial and skeletal muscle cell apoptosis. In addition, benfotiamine prevented the vascular accumulation of advanced glycation end products and the induction of pro-apoptotic caspase-3, while restoring proper expression of Nos3 and Akt in ischaemic muscles. The benefits of benfotiamine were nullified by dominant-negative PKB/Akt. In vitro, benfotiamine stimulated the proliferation of human EPCs, while inhibiting apoptosis induced by high glucose. In diabetic mice, the number of circulating EPCs was reduced, with the deficit being corrected by benfotiamine. CONCLUSIONS/INTERPRETATION: We have demonstrated, for the first time, that benfotiamine aids the post-ischaemic healing of diabetic animals via PKB/Akt-mediated potentiation of angiogenesis and inhibition of apoptosis. In addition, benfotiamine combats the diabetes-induced deficit in endothelial progenitor cells.

Animals↗

PubMed2005

Increased circulating levels of tissue kallikrein in systemic sclerosis correlate with microvascular involvement.

BACKGROUND: In systemic sclerosis (SSc) the lack of an angiogenic response to hypoxia may be due to inappropriate synthesis of angiogenic and angiostatic factors. Tissue kallikrein (t-kallikrein), regulating the kallikrein-kinin system and acting on the microcirculation, is a potent angiogenic agent, and kallistatin is its natural inhibitor. OBJECTIVE: To evaluate, in patients with SSc, t-kallikrein and kallistatin levels and their correlation with clinical features and measures of microvascular involvement. PATIENTS AND METHODS: Serum levels of t-kallikrein and kallistatin (ELISA) and t-kallikrein skin expression (immunohistochemistry) were studied in patients with SSc, and evaluated for subset (dSSc or lSSc), clinical and immunological features, and microvascular involvement (ulcers, telangiectasias, nailfold videocapillaroscopy). RESULTS: Circulating levels of t-kallikrein were higher in SSc than in controls (p<0.001). T-kallikrein did not differ between lSSc and dSSc, although it was higher in lSSc than in controls (p<0.001).T-kallikrein levels were higher in patients with early and active capillaroscopic pattern than in those with late pattern (p = 0.019 and 0.023). Patients with giant capillaries and capillary microhaemorrhages had higher t-kallikrein concentrations than patients with architectural derangement (p = 0.04). No differences in kallistatin levels were detected between patients with SSc and controls, or between lSSc and dSSc. In early SSc skin, the presence of t-kallikrein was found in endothelial and in perivascular inflammatory cells, while no staining in skin of advanced SSc was detected. CONCLUSION: T-kallikrein levels are increased in patients with SSc, particularly in lSSc, and are associated with early and active capillaroscopic patterns. T-kallikrein may play a part in SSc microvascular changes.

Adult↗

PubMed2005

Ex vivo gene transfer for improvement of transplanted pancreatic islet viability and function.

Human pancreatic islet transplantation has recently been shown to be successful in replacing pancreatic endocrine function into type 1 diabetic recipients. A major drawback, however, is the high amount of pancreatic ss cells required to render one single patient insulin-independent. Given the shortage of human beta cell donors, the majority of type 1 diabetic patients remain excluded from this therapeutic option. High number of islets are requested since substantial islet cell death and dysfunction occur within the first few hours and days after islet transplantation. Impaired vascularization of the engraft, the non-specific inflammatory reaction at the site of transplantation, together with the presence of active or memory autoimmune responses to islet autoantigens and allogeneic recognition contribute to apoptosis of ss cells and subsequent early graft function loss. This review will focus on ex vivo engineering of the islet graft by gene transfer to improve islet engraftment. An overview of currently used gene transfer techniques will be given and their potential will be discussed.

Animals↗

PubMed2004

Nerve growth factor supplementation reverses the impairment, induced by Type 1 diabetes, of hindlimb post-ischaemic recovery in mice.

AIMS/HYPOTHESIS: Type 1 diabetes increases the risk of peripheral ischaemia and impairs recovery once ischaemia occurs, probably because the healing process is hampered by diabetes-induced endothelial dysfunction. In normoglycaemic mice subjected to limb ischaemia, blockade of nerve growth factor (NGF) compromises reparative angiogenesis. In the present study, we evaluated if expressional alterations of endogenous NGF system components are associated with diabetes-related impairment in neovascularisation. In addition, we tested whether the correction of NGF liabilities benefits post-ischaemic healing of Type 1 diabetic animals. METHODS: Unilateral hindlimb ischaemia was produced in streptozotocin-induced Type 1 diabetic mice. Purified murine NGF (20 microg daily for 14 days) or PBS were injected into ischaemic adductors. Non-diabetic mice given PBS served as controls. Hindlimb blood flow was analysed sequentially for up to 14 days. At necroscopy, adductors were removed for quantification of microvessel density, endothelial cell apoptosis and NGF receptor expression. NGF content was determined by ELISA three days after ischaemia. In vitro, we tested whether NGF protects endothelial cells from apoptosis induced by high glucose and whether vascular endothelial growth factor-A (VEGF-A) is involved in this beneficial effect. RESULTS: Muscles removed from Type 1 diabetic mice showed reduced NGF content and up-regulation of the NGF p75 receptor. NGF supplementation promoted capillarisation and arteriogenesis, reduced apoptosis, and accelerated blood flow recovery. NGF stimulated VEGF-A production by human endothelial cells incubated in high-glucose medium and conferred resistance against high-glucose-induced apoptosis via a VEGF-A-mediated mechanism. CONCLUSIONS/INTERPRETATION: NGF protects endothelial cells from apoptosis induced by Type 1 diabetes and facilitates reparative neovascularisation. The findings may open up new therapeutic options for the treatment of diabetic complications.

Animals↗

PubMed2004

Nerve growth factor promotes reparative angiogenesis and inhibits endothelial apoptosis in cutaneous wounds of Type 1 diabetic mice.

AIMS/HYPOTHESIS: The neurotrophin nerve growth factor (NGF) is pro-angiogenic and facilitates wound repair. The present study was conducted to (i) assess the statement of NGF system components in diabetic wounds and (ii) evaluate whether NGF supplementation could prevent impairment of wound neoangiogenesis by diabetes. METHODS: Skin wounds were produced in the interscapular region of streptozotocin-induced diabetic mice. NGF (1 microg per day in PBS) or vehicle was applied onto the ulcers for 3 days after punching. Non-diabetic mice were used as controls. RESULTS: In wounds of untreated diabetic mice, endogenous levels of immunoreactive NGF were lower than those in wounds of non-diabetic mice ( p<0.01). Immunohistochemical analysis showed down-regulation of tyrosine kinase receptor-A (TrkA) and up-regulation of p75 receptor in granulation tissue microvasculature. Local NFG administration prevented diabetes-induced expressional alterations, enhanced reparative capillarisation ( p<0.01), and accelerated wound closure ( p<0.01). This was associated with a three-fold increase in endothelial cell proliferation ( p<0.01), while apoptosis was reduced by 50% ( p<0.05). Quantitative RT-PCR documented a 5.5-fold increase in the expression of vascular endothelial growth factor-A (VEGF-A) by exogenous NGF in diabetic tissues ( p<0.01). In in vitro preparations of human endothelial cells from derma, NGF increased the release of immunoreactive VEGF-A, and reduced high-glucose-induced apoptosis ( p<0.05), the latter effect being inhibited by a VEGF-A receptor-2 antagonist. CONCLUSIONS/INTERPRETATION: Diabetic ulcers display distinct alterations in reparative angiogenesis and in the expression of NGF and its receptors. NGF supplementation corrects endogenous liabilities, facilitates vascular regeneration, and suppresses endothelial apoptosis seemingly via VEGF-A. Our findings unravel new mechanisms responsible for NGF reparative action.

Administration, Cutaneous↗

PubMed2004

Angiogenesis therapy with human tissue kallikrein for the treatment of ischemic diseases.

Angiogenesis is essential for the repair of wounds and tissues damaged by ischemia. The regenerative process is tightly regulated by master angiogenic factors, cytokines and the downstream mediator NO. In addition, modulators of vascular growth, such as COX-2-generated prostanoids, contribute to the process by stabilizing the hypoxia-inducible factor and stimulating the expression of VEGF. Recently, we discovered that human tissue kallikrein, a member of the serine proteinase superfamily, possesses potent angiogenic effects. It has been categorized as a pleiotropic angiogenic agent acting via enzymatic cleavage of kininogen and subsequent release of kinin peptides. Kinins bind G-protein coupled receptors, subtype B1 and B2, and exert proliferative effects on endothelial cells via an IP3K-Akt-NO mediated mechanism independent of VEGF. In addition, kinins stimulate the release of angiogenic prostacyclin. Gene transfer of human tissue kallikrein rescues ischemic tissues in otherwise normal mice, as well as in hypertensive or diabetic animals. In addition, prophylactic gene delivery of tissue kallikrein to diabetic skeletal muscles prevents the development of microangiopathy and stimulates collateralization, thus protecting from the consequences of supervening arterial occlusion.

Animals↗

PubMed2001

Giorgio Olivetti.

Explore the source record for details and available documents.

Cardiovascular Diseases↗

PubMed2001

Local delivery of human tissue kallikrein gene accelerates spontaneous angiogenesis in mouse model of hindlimb ischemia.

BACKGROUND: Human tissue kallikrein (HK) releases kinins from kininogen. We investigated whether adenovirus-mediated HK gene delivery is angiogenic in the context of ischemia. METHODS AND RESULTS: Hindlimb ischemia, caused by femoral artery excision, increased muscular capillary density (P:<0.001) and induced the expression of kinin B(1) receptor gene (P:<0.05). Pharmacological blockade of B(1) receptors blunted ischemia-induced angiogenesis (P:<0.01), whereas kinin B(2) receptor antagonism was ineffective. Intramuscular delivery of adenovirus containing the HK gene (Ad. CMV-cHK) enhanced the increase in capillary density caused by ischemia (969+/-32 versus 541+/-18 capillaries/mm(2) for control, P:<0.001), accelerated blood flow recovery (P:<0.01), and preserved energetic charge of ischemic muscle (P:<0.01). Chronic blockade of kinin B(1) or B(2) receptors prevented HK-induced angiogenesis. CONCLUSIONS: HK gene delivery enhances the native angiogenic response to ischemia. Angiogenesis gene therapy with HK might be applicable to peripheral occlusive vascular disease.

Adenoviridae↗

PubMed2001

Targeting kinin receptors for the treatment of tissue ischaemia.

Kinins, the biological end-products of the kallikrein-kininogen system, influence many aspects of the cellular function. Interest in this peptidergic system has been renewed recently by the discovery that kinins exert cardiovascular protective effects and promote post-ischaemic recovery by stimulating vascular growth. Pharmacological and genetic studies indicate that induction of kallikrein and kinin receptors by ischaemia is functionally relevant in the natural host response that permits perfusion recovery and tissue healing. Furthermore, potentiation of the generation of kinins by continuous supply of tissue kallikrein promotes reparative angiogenesis through stimulation of the release of nitric oxide and prostaglandins. Strategies that activate kinin receptors might be applicable to the treatment of occlusive vascular disease, whereas kinin receptor antagonists could represent therapeutic reagents against pathological angiogenesis in cancer and chronic inflammatory conditions.

Animals↗

PubMed2001

Role of calcitonin gene-related peptide and kinins in post-ischemic intestinal reperfusion.

The involvement of kinins, calcitonin gene-related peptide (CGRP), and tachykinins during mesenteric post-ischemic reperfusion was studied in anesthetized rats by using antagonists for bradykinin (BK) B1, BK B2, CGRP1, or tachykinin NK1 receptor, or by capsaicin-induced desensitization. B1, B2, or CGRP1 receptor antagonists or desensitization attenuated the transient hypotension and plasma protein and leukocyte infiltration of intestinal wall observed during post-ischemic reperfusion. These effects were abolished by the combination of B2 and CGRP1 blockade as well as by B2 antagonism in capsaicinized rats, while NK1 blockade was ineffective. Our results suggest that kinins and CGRP contribute to systemic vasodilatation and microvascular leakage during mesenteric reperfusion. Pharmacological blockade of these systems could help preventing hypotension and intestinal injury consequent to reperfusion.

Animals↗

PubMed2001

Angiogenesis gene therapy to rescue ischaemic tissues: achievements and future directions.

Ischaemic diseases are characterized by an impaired supply of blood resulting from narrowed or blocked arteries that starve tissues of needed nutrients and oxygen. Coronary-atherosclerosis induced myocardial infarction is one of the leading causes of mortality in developed countries. Ischaemic disease also affects the lower extremities. Considerable advances in both surgical bypassing and percutaneous revascularization techniques have been reached. However, many patients cannot benefit from these therapies because of the extension of arterial occlusion and/or microcirculation impairment. Consequently, the need for alternative therapeutic strategies is compelling. An innovative approach consists of stimulating collateral vessel growth, a natural host defence response that intervenes upon occurrence of critical reduction in tissue perfusion (Isner & Asahara, 1999). This review will debate the relevance of therapeutic angiogenesis for promotion of tissue repair. The following issues will receive attention: (a) vascular growth patterns, (b) delivery systems for angiogenesis gene transfer, (c) achievements of therapeutic angiogenesis in myocardial and peripheral ischaemia, and (d) future directions to improve effectiveness and safety of vascular gene therapy.

Animals↗

PubMed2001

Renal phenotype of low kallikrein rats.

BACKGROUND: Renal kallikrein has been linked with inheritance of arterial hypertension and with sensitivity to drug nephrotoxicity. Identification of a cause--effect relationship between low kallikrein and intermediate phenotypes has been hampered by the lack of adequate animal models. METHODS: Kallikrein was measured in tissues obtained from rats inbred for low urinary kallikrein excretion (LKR) and wild-type controls. Blood pressure and indices of myocardial contractility were recorded via an intraventricular cannula connected to a transducer. The functional relevance of endogenous angiotensin II (Ang II) in LKR was explored by determining the effect of Ang II subtype 1 (AT(1)) receptor blockade on glomerular filtration rate, renal blood flow, and urinary sodium excretion. In addition, sensitivity to gentamycin-induced nephrotoxicity was evaluated. RESULTS: Kallikrein activity was reduced by 60% in the kidney of LKR (P < 0.01), whereas it was increased in the heart (P < 0.05) and was unaltered in the pancreas, liver, and salivary glands. Heart rate and myocardial contractility were reduced, and the mean blood pressure (MBP) was increased in LKR as compared with controls (P < 0.05). LKR exhibited polydipsia, polyuria, glomerular hyperfiltration, and reduced fractional sodium excretion under basal conditions and impaired renal vasodilation in response to volume expansion. These functional alterations were significantly attenuated by AT(1) receptor blockade. Gentamycin reduced the glomerular filtration rate in LKR, but not in controls. CONCLUSIONS: In LKR, unopposed activity of Ang II appears to be responsible for increased glomerular hydrostatic pressure and augmented tubular reabsorption. Balance between the kallikrein-kinin and renin-angiotensin systems is essential for normal renal function.

Angiotensin Receptor Antagonists↗

PubMed2001

Rescue of impaired angiogenesis in spontaneously hypertensive rats by intramuscular human tissue kallikrein gene transfer.

Angiogenesis represents a compensatory response targeted to preserve the integrity of tissues subjected to ischemia. The aim of the present study was to examine whether reparative angiogenesis is impaired in spontaneously hypertensive rats (SHR), as a function of progression of hypertension. In addition, the potential of gene therapy with human tissue kallikrein (HK) in revascularization was challenged in SHR and normotensive Wistar-Kyoto rats (WKY) that underwent excision of the left femoral artery. Expression of vascular endothelial growth factor and HK was upregulated in ischemic hindlimb of WKY but not of SHR. Capillary density was increased in ischemic adductor muscle of WKY (from 266+/-20 to 633+/-73 capillaries/mm(2) at 28 days, P<0.001), whereas it remained unchanged in SHR (from 276+/-20 to 354+/-48 capillaries/mm(2), P=NS), thus compromising perfusion recovery as indicated by reduced plantar blood flow ratio (0.61+/-0.08 versus 0.92+/-0.07 in WKY at 28 days, P<0.05). In separate experiments, saline or 5x10(9) pfu adenovirus containing the HK gene (Ad.CMV-cHK) or the beta-galactosidase gene (Ad.CMV-LacZ) was injected intramuscularly at 7 days after the induction of ischemia. Ad.CMV-cHK augmented capillary density and accelerated hemodynamic recovery in both strains, but these effects were more pronounced in SHR (P<0.01). Our results indicate that native angiogenic response to ischemia is impaired in SHR, possibly as a result of defective modulation of endothelial cell mitogens. Supplementation with kallikrein, one of the growth factors found to be deficient in SHR, restores physiological angiogenic response utilitarian for tissue healing. Our discoveries may have important implications in vascular medicine for therapeutic benefit.

Animals↗

PubMed2000

Fetal ontogeny and role of metanephric bradykinin B2 receptors.

Previous studies in rats have shown that blockade of bradykinin B2 receptors (B2R) in combination with a high-salt intake during gestation result in poor postnatal survival and long-term hypertension in the offspring. In this study, we examined the fetal ontogeny of B2R and determined the consequences of gestational B2R blockade and high salt on kidney development. B2R gene expression is induced on embryonic day (E16) of fetal metanephrogenesis and remains sustained until term. The earliest expression of the B2R protein is observed on apical membranes of ureteric bud branches and in capillary loop stage glomeruli. By the end of gestation, B2R becomes restricted to more-differentiated tubules in the deep cortex and medulla. Pairs of rats on normal (0.12 mmol/g) or high (0.84 mmol/g) salt diets were mated at 14 weeks of age. The B2R antagonist, Icatibant (previously known as Hoe-140) (300 nmol/kg per day) or saline (vehicle) was infused intraperitoneally during gestation via osmotic minipumps. Fetuses were examined on E20 (n=27-36 per group). No significant differences in litter size or body weight were observed among the groups. Combined high-salt and Icatibant treatment caused aberrant fetal renal development characterized by tubular dysgenesis, widened stromal mesenchyme, and glomerular cysts. The dysgenetic tubules stained positively for the distal nephron lectin, Dolichos biflorus, and exhibited enhanced Bax expression and apoptosis. Renal microvascular development, the number of mature glomeruli, and percentage of proliferating glomerular cells were not affected. Gestational Icatibant or high salt alone had no deleterious effects on fetal nephrogenesis. We conclude that gestational blockade of the kallikrein-kinin system impairs fetal nephrogenesis if combined with an intrauterine stressor such as high-salt intake. B2R may play a protective role during segmental nephron differentiation.

Abnormalities, Drug-Induced↗

PubMed2000

Studies of the cardiovascular effects of nociceptin and related peptides.

Nociceptin, a novel opioid peptide, and its ORL(1) receptor share structural similarities with other opioid ligands and receptors. Although NC exerts evident cardiovascular effects at a central and peripheral level, its role in homeostatic mechanisms and disease states are just beginning to be understood, as only recently selective receptor antagonists became available. In this review, some of the new observations regarding the cardiovascular actions of NC, related peptides and newly synthesized receptor antagonists are discussed.

Animals↗

PubMed2000

Participation of kinins in the captopril-induced inhibition of intimal hyperplasia caused by interruption of carotid blood flow in the mouse.

1. In the rat balloon injury model, angiotensin-converting enzyme (ACE) inhibitors prevent vascular remodelling by inhibiting angiotensin II generation and kinin breakdown. We investigated if ACE inhibition also prevents the structural vascular responses to disruption of carotid artery blood flow and if kinin potentiation plays a role in such a protection. 2. Morphometric analysis of the structural alterations caused by ligation of the left carotid artery was performed 14 days after surgery in J129Sv wild-type mice (B(2)(+/+)) drinking normal tap water or water containing captopril (120 mg kg(-1) per day). In addition, the effect of captopril on vascular remodelling was tested in B(2)(+/+) given the bradykinin (BK) B(1) receptor antagonist des-Arg(9)-[Leu(8)]-BK (DALBK, 50 nmol kg(-1) per day, intraperitoneally) or the BK B(2) receptor antagonist D-Arg, [Hyp(3),Thi(5)D-Tic(7),Oic(8)]-BK (icatibant, 1 micromol kg(-1) per day, intraperitoneally), and in B(2) receptor gene knockout mice (B(2)(-/-)). 3. Interruption of blood flow resulted in carotid artery intimal hyperplasia and media thickening in untreated B(2)(+/+), these responses being partially suppressed by captopril. The inhibition of intimal thickening exerted by captopril was reduced in B(2)(+/+) given DALBK or icatibant (P<0.05 for both comparisons) as well as in B(2)(-/-) (P<0.05). Neither antagonism of kinin receptors nor disruption of the B(2) receptor gene altered the suppressive effect of captopril on media thickening. The protection of vascular wall structure was independent of the reduction in blood pressure by captopril. 4. These results demonstrate that kinins participate in the inhibitory effect of captopril on intimal hyperplasia via B(1) and B(2) receptor signalling. Our findings may have important implications in treating vascular remodelling evoked by altered shear stress conditions.

Angiotensin-Converting Enzyme Inhibitors↗