Begoña Cuevas

Peripheral nerve regeneration by bone marrow stromal cells

Abstract Adult bone marrow contains stem cells that have attracted interest through their possible use for cell therapy in neurological diseases. Bone marrow stromal cells (MSCs) were harvested from donor adult rats, cultured and pre-labeled with bromodeoxyuridine (BrdU) previously to be injected in the distal stump of transected sciatic nerve of the rats. Distal nerve stump of control rats received culture medium solution. MSCs-treated rats exhibit significant improvement on walking track test at days 18 and 33 compared to controls. Dual immunofluorescence labeling shows that BrdU reactive cells survive in the injected area of transected sciatic nerve at least 33 days after implantation, and almost 5% of BrdU cells express Schwann cell-like phenotype (S100 immunoreactivity). Because MSCs injected in a lesioned peripheral nerve can survive, migrate, differentiate in Schwann cells, and promote functional recovery, they may be an important source for cellular therapy in several neurological diseases.

Angiogenic capacity of human omental stem cells

Abstract Objectives: The goals of the present study are to obtain, expand and characterize a stem cell population from human omentum and to evaluate its in vivo angiogenic capacities. Methods: Human omental CD34+ cells were obtained from samples of human omentum by density gradient centrifugation in Ficoll. Proliferative pattern, marker expression (by flow cytometry) and angiogenic growth factor synthesis by omental cell cultures were determined. In vivo angiogenic capacity of the cells was evaluated in rats. Results: Omental stem cells showed a high rate of proliferation (Ki67 staining), expressed CD34 marker and synthesized bFGF and VEGF. When implanted in rats, omental cells promoted neovascularization. Human omental cells were localized in rat tissue, mainly forming the endothelium of neo-vessels. Implantation of omental cells also facilitated angiogenesis of rat origin. Conclusion: CD34+ cell population of human omentum could be responsible for the clinical benefit of omental transplantation by promoting angiogenesis and synthesizing angiogenic growth factors to facilitate revascularization of injured tissue.

Vardenafil enhances clitoral and vaginal blood flow responses to pelvic nerve stimulation in female dogs

The relaxation of the smooth muscle in the vagina and clitoris and the increase of blood flow into these organs is thought to be essential in the female sexual response. Vardenafil is a type 5 phosphodiesterase (PDE5) inhibitor that potentiates the nitric oxide (NO)/cGMP pathway facilitating penile smooth muscle relaxation and improving penile erection in men. Although the potentiation of the NO/cGMP pathway through PDE5 inhibitors can clearly enhance blood flow into the penis and is used in the therapy of male sexual dysfunction, there is controversy about the efficacy of these agents in improving female sexual function. The aim of this work was to evaluate the effects of vardenafil on the increase of blood flow into the vagina and clitoris induced by pelvic nerve electrical stimulation (PNES) in a female dog model. Application of PNES produced consistent and frequency-related increased blood flow into the vagina and clitoris of anesthetized female dogs. The magnitude and duration of the blood flow responses to PNES were variable among the different animals but remained stable over time within the same animal. The intravenous administration of vardenafil (1 mg/kg) significantly potentiated the increases in blood flow produced by PNES into the vagina (381.4 and 206.2% of control response at 5 and 10 Hz, respectively, P<0.01, n=6) and clitoris (379.4 and 238.5% of control response at 5 and 10 Hz, respectively, P<0.01, n=6) 20 min after administration. The significant enhancement of PNES-induced responses was maintained 50 min (224.5 and 181.0%, P<0.01 in vagina; 294.8 and 258.9%, P<0.05 in clitoris) and 80 min after vardenafil administration (209.5 and 156.9%, P<0.05 in vagina; 268.9 and 194.9%, P<0.05 in clitoris). Here we present a feasible model for research into female sexual function. Our results show that vardenafil effectively potentiates the blood flow responses to PNES in the genitalia of female dogs. These results emphasize the role of the NO/cGMP pathway in the local vasodilatory response in female sexual organs and provide a rationale for testing PDE5 inhibitors, such as vardenafil, as a treatment for certain forms of female sexual dysfunction.

Single topical application of human recombinant basic fibroblast growth factor (rbfgf) promotes neovascularization in rat cerebral cortex

Basic fibroblast growth factor (bFGF) is a morphogenic, chemotactic, mitogenic, and angiogenic peptide found within the central nervous system (CNS) with potent neurotrophic effects. A potential role in ischemia-induced vascular growth has been suggested for bFGF. In this study, we show that single, topical administration of human recombinant basic fibroblast growth factor (rbFGF) in the rat cerebral cortex promotes capillary overgrowth and might mimic the angiogenic response observed after brain ischemia. The implication of this finding for the therapeutic use of basic FGF in angiogenic therapy is discussed.

Inhibition of vascular endothelial growth factor (VEGF)-induced endothelial proliferation, arterial relaxation, vascular permeability and angiogenesis by dobesilate.

Vascular endothelial growth factor (VEGF) is a key factor in angiogenesis and vascular permeability which is associated with many pathological processes. 2,5-hydroxybenzene sulfonate (DHBS; dobesilate) is a small molecule with anti-angiogenic activity that has been described as an inhibitor of fibroblast growth factors (FGF). The aim of the present study was to evaluate the effects of DHBS on VEGF-induced actions. The effects of DHBS were evaluated on VEGF-induced proliferation in human umbilical vein endothelial cells (HUVEC) and rat aorta relaxation, as well as on in vivo VEGF-induced skin vascular permeability and neovascularization in rats. DHBS at 50 and 100 μM concentration significantly inhibited the proliferation of HUVEC induced by VEGF (10 ng/ml), without significantly affecting HUVEC proliferation in the absence of VEGF. Rapid VEGF-induced activation of Akt in HUVEC was also prevented by DHBS (100 μM). Additionally, DHBS (2 μM) specifically inhibited the relaxation of rat aorta induced by VEGF (0.1 to 30 ng/ml), but not endothelium-dependent relaxation to acetylcholine (1 nM to 10 μM). The in vivo enhancement of vascular permeability caused by VEGF injection (50 μl at 10 ng/ml) in rat skin was also inhibited by DHBS co-administration (200 μM) (74.8±3.8% inhibition of dye extravasation). Administration of DHBS (200 mg/kg/day; i.p.) also reduced VEGF-induced angiogenesis in vivo. DHBS inhibits main responses elicited in vitro and in vivo by VEGF. As a dual antagonist of VEGF and FGF activities, DHBS could be of therapeutic interest in the treatment of diseases related to VEGF/FGF overproduction and excessive angiogenesis.

Osteopromotion for cranioplasty: An experimental study in rats using acidic fibroblast growth factor

BACKGROUND Many growth factors influence the bone healing cascade. Furthermore, the occasional failure of bone repair may in part be due to perturbation in the activation of local growth factors. Local activation of fibroblast growth factors (FGFs) at the fracture site may serve to increase neovascularization, and induce early granulation formation that can affect bone healing. METHODS We have performed a rat parietal (6 x 3 mm) critical size defect (CSD). Human recombinant acidic fibroblast growth factor (hraFGF) imbided in agarose was topically administered at the craniectomy site. Control animals received agarose alone in the same manner. Three weeks after surgery, osteopromotion was histologically evaluated. RESULTS hraFGF-treated animals show a continuous bridge of regenerated bone extending from one edge of the defect to the other. None of the parietal defects that had been treated with agarose contained new bone in the central portion. CONCLUSION Our results suggest an important role of FGFs to promote large cranioplasty repair and support the use of these proteins as an alternative choice for bone grafts and bone substitutes.

Nebivolol Dilates Human Penile Arteries and Reverses Erectile Dysfunction in Diabetic Rats through Enhancement of Nitric Oxide Signaling

INTRODUCTION Traditional beta-blockers have sometimes been associated with erectile dysfunction (ED). Nebivolol is a cardioselective β(1)-adrenoceptor antagonist that promotes vasodilation through a nitric oxide (NO)-dependent mechanism. AIM We evaluated the effects of nebivolol on the NO/cyclic guanosine monophosphate (cGMP) signaling pathway, on erectile function and dysfunction, and in human penile vascular tissues. METHODS Erectile response to cavernosal nerve electrical stimulation in control and diabetes-induced ED rats were evaluated, along with serum nitrite/nitrate (NOx) concentration and plasma/tissue cGMP levels. Endothelium-dependent and sildenafil-induced relaxation of isolated human corpus cavernosum (HCC) and human penile resistance arteries (HPRA) were also determined. MAIN OUTCOME MEASURES The effects of nebivolol on erectile function and dysfunction and on NO/cGMP-mediated responses. RESULTS Treatment with nebivolol significantly potentiated erectile response in control rats, regardless of its effects on blood pressure. Nebivolol increased NOx and plasma cGMP by 3-fold and 2.75-fold, respectively, and significantly augmented the elevation of plasma cGMP produced by sildenafil. Nebivolol enhanced endothelium-dependent and sildenafil-induced relaxations of HCC tissue, and produced endothelium-dependent vasodilation of HPRA. Nebivolol, but not atenolol, significantly improved erectile response in diabetic rats (51.6%, 53.2%, and 87.1% of response at 3 Hz in nondiabetic rats, for vehicle-treated, atenolol-treated, and nebivolol-treated diabetic rats, respectively); after sildenafil administration, ED was completely reversed in nebivolol-treated diabetic rats (69.6% and 112% for diabetic rats treated with sildenafil and nebivolol plus sildenafil, respectively). Accordingly, nebivolol restored systemic NOx levels and cGMP content in penile tissue from these animals. CONCLUSIONS Nebivolol in vivo activated the NO/cGMP pathway, enhanced erectile response and reversed ED in diabetic rats. Moreover, nebivolol in vitro potentiated NO/cGMP-mediated relaxation of human erectile tissues. These effects may account for the low incidence of ED in nebivolol-treated hypertensive patients. Nebivolol therefore may have utility in the treatment of ED, particularly ED associated with diabetes.

Inhibition of intra-tumoral angiogenesis and glioma growth by the fibroblast growth factor inhibitor 1,3,6-naphthalenetrisulfonate

1,3,6-naphthalenetrisulfonate (NTS) can inhibit the proliferation in vitro of cells of various origin including glioma. We have studied the effects of NTS on intra-tumoral angiogenesis and tumor growth in the rabbit cornea after implantation of C6 rat glioma cells. It was found that neovascularization and glioma growth were abolished by topical administration of NTS. This effect could be mediated by both induction of programmed cell death and inhibition of growth, in endothelium and in tumor cells, most likely as a consequence of the disruption of the autocrine and paracrine effects of FGF released from endothelial and tumor cells. The results suggest that NTS is a promising candidate to lead the development of new angiogenesis inhibitors for the treatment of cancer and other diseases whose progression is dependent upon the development of new blood vessels.

Antidepressant-Induced Inhibition of Genital Vascular Responses Is Reversed by Vardenafil in Female Rabbits

INTRODUCTION Administration of serotonin reuptake inhibitors (SRI) or serotonin and norepinephrine reuptake inhibitors (SNRI) relieves depressive symptoms but may cause sexual dysfunction in women and men. AIM The aim of the present study was to evaluate the effects of the phosphodiesterase type 5 (PDE5) inhibitor, vardenafil, on inhibition of genital vascular responses (GVR) induced by SRI or SNRI administration in female rabbits. METHODS Vaginal and clitoral vasodilatory responses to pelvic nerve electrical stimulation were measured by laser Doppler flow needle probes. RESULTS GVR were significantly potentiated by vardenafil even at the low dose of 0.1 mg/kg, in clitoris and vagina (181 +/- 22% and 180 +/- 31% of control, in vagina and clitoris, respectively, at 8 Hz). The selective SRI, paroxetine (5 mg/kg), significantly inhibited GVR in female rabbits (54 +/- 5% and 48 +/- 6% of control). GVR were also significantly inhibited by the SNRIs, venlafaxine (5 mg/kg) (57 +/- 3% and 32 +/- 11%) and duloxetine (1 mg/kg) (40 +/- 7% and 28 +/- 5%). Treatment with vardenafil (0.1 and 0.3 mg/kg) completely reversed the inhibition of GVR induced by paroxetine, venlafaxine, or duloxetine. CONCLUSIONS Potentiation of the nitric oxide (NO) pathway by vardenafil improves vascular sexual responses in female rabbits and overcomes the inhibitory effects of acutely administered antidepressants on GVR, irrespective of the underlying pathophysiologic mechanism, i.e., disruption of the NO pathway or enhancement of alpha-adrenergic mechanisms. PDE5 inhibition may represent a reasonable approach to treat SRI- or SRNI-induced female sexual dysfunction, in particular, arousal disorders.

Nitrergic innervation of rat brain vasculature

Nitric oxide synthase has been located in the brain, in several neuronal populations which appear in close contact either with the microvasculature or the external surface of extracerebral vessels. These data provide structural support to the pharmacological evidences of the nitric oxide participation in the regulation of vascular tone of brain blood vessels.