Alessandra Micera

Publications (43)140.22 Total impact

• Source

  Available from: Bijorn Omar Balzamino

  Article: Chronic nerve growth factor exposure increases apoptosis in a model of in vitro induced conjunctival myofibroblasts.

  Alessandra Micera, Ilaria Puxeddu, Bijorn Omar Balzamino, Stefano Bonini, Francesca Levi-Schaffer
  [show abstract] [hide abstract]
  ABSTRACT: In the conjunctiva, repeated or prolonged exposure to injury leads to tissue remodeling and fibrosis associated with dryness, lost of corneal transparency and defect of ocular function. At the site of injury, fibroblasts (FB) migrate and differentiate into myofibroblasts (myoFB), contributing to the healing process together with other cell types, cytokines and growth factors. While the physiological deletion of MyoFB is necessary to successfully end the healing process, myoFB prolonged survival characterizes the pathological process of fibrosis. The reason for myoFB persistence is poorly understood. Nerve Growth Factor (NGF), often increased in inflamed stromal conjunctiva, may represent an important molecule both in many inflammatory processes characterized by tissue remodeling and in promoting wound-healing and well-balanced repair in humans. NGF effects are mediated by the specific expression of the NGF neurotrophic tyrosine kinase receptor type 1 (trkA(NGFR)) and/or the pan-neurotrophin glycoprotein receptor (p75(NTR)). Therefore, a conjunctival myoFB model (TGFβ1-induced myoFB) was developed and characterized for cell viability/proliferation as well as αSMA, p75(NTR) and trkA(NGFR) expression. MyoFB were exposed to acute and chronic NGF treatment and examined for their p75(NTR)/trkA(NGFR), αSMA/TGFβ1 expression, and apoptosis. Both NGF treatments significantly increased the expression of p75(NTR), associated with a deregulation of both αSMA/TGFβ1 genes. Acute and chronic NGF exposures induced apoptosis in p75(NTR) expressing myoFB, an effect counteracted by the specific trkA(NGFR) and/or p75(NTR) inhibitors. Focused single p75(NTR) and double trkA(NGFR)/p75(NTR) knocking-down experiments highlighted the role of p75(NTR) in NGF-induced apoptosis. Our current data indicate that NGF is able to trigger in vitro myoFB apoptosis, mainly via p75(NTR). The trkA(NGFR)/p75(NTR) ratio in favor of p75(NTR) characterizes this process. Due to the lack of effective pharmacological agents for balanced tissue repairs, these new findings suggest that NGF might be a suitable therapeutic tool in conditions with impaired tissue healing.
  PLoS ONE 01/2012; 7(10):e47316. · 4.09 Impact Factor
• Source

  Available from: Sergio Bonini

  Article: TLR4 and TLR9 Expression in Different Phenotypes of Rhinitis.

  Maria Lauriello, Alessandra Micera, Paola Muzi, Lino Di Rienzo Businco, Sergio Bonini
  [show abstract] [hide abstract]
  ABSTRACT: Background. Toll-like receptors (TLRs) represent a family of evolutionarily conserved proteins, that represent a fundamental link between innate and adaptive immune responses. Aim. The purpose of this study was to investigate the expression of TLR4 and TLR9 in the normal nasal mucosa and in the mucosa of subjects with different phenotypes of rhinitis. Methodology. A confocal analysis of TLR4 and TLR9 (co)expression was carried out on biopsies from the inferior turbinate obtained from 4 patients affected by persistent allergic rhinitis, 8 patients with chronic rhino-sinusitis, and 6 patients with vasomotor rhinitis The results were compared with those of specimens obtained from 4 subjects undergoing nasal surgery, but with signs of nasal inflammation. Results. TLR4 and TLR9 were expressed in the healthy nasal mucosa; TLR4 and TLR9 expression was significantly decreased in allergic rhinitis. TLR4 was over expressed in the epithelium of chronic rhino-sinusitis. Both TLRs were co-expressed in the sub-epithelial infiltrate of chronic and vasomotor rhinitis, even though this expression was higher in the former compared with the latter. Conclusions. This study indicates that TLR4 and TLR9 show a different pattern of expression in different phenotypes of rhinitis, possibly related to the type and severity of the disease.
  International Journal of Otolaryngology 01/2012; 2012:925164.
• Article: Alterations of tear neuromediators in dry eye disease.

  Alessandro Lambiase, Alessandra Micera, Marta Sacchetti, Magdalena Cortes, Flavio Mantelli, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: To evaluate tear levels of neuromediators in patients with dry eye disease and to identify statistical correlations with the clinical findings. Nineteen patients with dry eye disease (Sjögren syndrome, n = 5 patients; non-Sjögren syndrome, n = 10; and ocular cicatricial pemphigoid, n = 4) and 12 healthy volunteers were enrolled. The eyes of all participants were evaluated by slitlamp examination, Schirmer testing, fluorescein staining, and tear film break-up time. Grading of dry eye severity was recorded. Tear samples were collected, and substance P, calcitonin gene-related peptide (CGRP), neuropeptide Y (NPY), vasoactive intestinal peptide, and nerve growth factor (NGF) concentrations were evaluated by enzyme-linked immunoassay and correlated with the clinical findings. Nerve growth factor tear levels were significantly increased in participants with dry eye disease; CGRP and NPY concentrations were significantly decreased when compared with those in healthy participants. Dry eye severity showed a direct correlation with NGF and an inverse correlation with CGRP and NPY tear levels. Nerve growth factor tear levels showed a direct correlation with conjunctival hyperemia and fluorescein staining results, CGRP directly correlated with Schirmer test values, and NPY inversely correlated with tear film break-up time. Subgroup analysis showed that CGRP and NPY but not NGF were changed in autoimmune (ie, Sjögren syndrome and ocular cicatricial pemphigoid) dry eye disease. The decreased tear levels of NPY and CGRP in dry eye disease are related to impaired lacrimal function, and tear levels of NGF are more closely related to corneal epithelial damage. Our findings suggest that NPY, CGRP, and NGF could become useful markers of dry eye severity.
  Archives of ophthalmology 08/2011; 129(8):981-6. · 3.86 Impact Factor
• Article: Toll-like receptors in ocular surface diseases: overview and new findings.

  Alessandro Lambiase, Alessandra Micera, Marta Sacchetti, Flavio Mantelli, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: The ocular surface is the first line of defence in the eye against environmental microbes. The ocular innate immune system consists of a combination of anatomical, mechanical and immunological defence mechanisms. TLRs (Toll-like receptors), widely expressed by the ocular surface, are able to recognize microbial pathogens and to trigger the earliest immune response leading to inflammation. Increasing evidence highlights the crucial role of TLRs in regulating innate immune responses during ocular surface infective and non-infective inflammatory conditions. In addition, recent observations have shown that TLRs modulate the adaptive immune response, also playing an important role in ocular autoimmune and allergic diseases. One of the main goals of ocular surface treatment is to control the inflammatory reaction in order to preserve corneal integrity and transparency. Recent experimental evidence has shown that specific modulation of TLR pathways induces an improvement in several ocular inflammatory conditions, such as allergic conjunctivitis, suggesting new therapeutic anti-inflammatory strategies. The purpose of the present review is to summarize the current knowledge of TLRs at the ocular surface and to propose them as potential targets of therapy for ocular inflammatory conditions.
  Clinical Science 05/2011; 120(10):441-50. · 4.61 Impact Factor
• Source

  Available from: Sergio Bonini

  Article: Tear levels of neuropeptides increase after specific allergen challenge in allergic conjunctivitis.

  Marta Sacchetti, Alessandra Micera, Alessandro Lambiase, Stefania Speranza, Flavio Mantelli, Girolamo Petrachi, Sergio Bonini, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: Growing evidence is showing a role of neurogenic inflammation in allergic reactions, with sensory and autonomic nerve fibers releasing neuromediators, which may actively participate in the allergic inflammatory cascade. Although the cornea is the most densely innervated tissue of the human body, little is known on the role of neuromediators at the ocular surface. In this study, we aimed at evaluating the role of substance P (SP), calcitonine gene related peptide (CGRP), neuropeptide Y (NPY) and vasoactive intestinal peptide (VIP) in allergic reactions of the ocular surface. Fifteen patients with allergic conjunctivitis (6 female, 9 male, mean age 30±8 years) in non-active phase, and 10 age-matched healthy subjects were included in this study. The conjunctival provocation test (CPT) with allergen was performed in all allergic patients and in 5 healthy subjects. Tear samples were collected and the tear content of VIP, NPY, CGRP, and SP was measured by ELISA at baseline and after CPT. The Mann-Whitney U-test and Wilcoxon test were used to compare neuromediator tear levels. No significant differences in neuropeptide tear levels were observed between healthy and allergic patients in non-active phase. CPT induced conjunctival hyperemia and itching in all allergic patients, while no reaction was observed in the control eyes and in healthy subjects. In allergic patients SP, CGRP, and VIP, but not NPY, were significantly higher after CPT as compared to baseline (SP: 3.9±1.3 ng/ml versus 5.8±1.1 ng/ml, p=0.011; CGRP: 5.5±2.3 ng/ml versus 7.3±2.7 ng/ml; p=0.002; VIP: 4±0.9 ng/ml versus 5.1±1.5 ng/ml, p=0.007). No significant changes were observed in the control eyes of allergic patients challenged with diluent and in healthy subjects after allergen provocation. Locally-released neuromediators may participate in modulating the allergic response of the ocular surface.
  Molecular vision 01/2011; 17:47-52. · 2.20 Impact Factor
• Article: Neurogenic inflammation of the ocular surface.

  Flavio Mantelli, Alessandra Micera, Marta Sacchetti, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: The purpose of the present review is to describe the involvement of neurogenic inflammation in ocular surface diseases and to outline recent advances in understanding the complex interaction of neuromediators and inflammatory cells in the conjunctiva. Evidence indicates that all clinical forms of ocular surface disease, including allergic and autoimmune, share the involvement of a neurogenic inflammation triggered by nociceptive stimuli and resulting in plasma extravasation and local activation of immune cells. Players of this complex mechanism are neuropeptides, including substance P, calcitonine gene-related peptide, neuropeptide Y, and vasoactive intestinal peptide. These neuropeptides are, however, only a minor component of a complex neuroimmune cross-talk capable of modulating local inflammation. As already shown in other mucosal surfaces, neurogenic inflammation and innate immunity may work together to protect the ocular surface. In this review, we present the most recent advances in the understanding of the roles played by nerve endings and neuropeptides in local inflammatory processes of the ocular surface, in order to better clarify their function in physiologic and pathologic conditions and to open new research, diagnostic, and therapeutic perspectives.
  Current Opinion in Allergy and Clinical Immunology 10/2010; 10(5):498-504. · 4.11 Impact Factor
• Article: Multiple action agents and the eye: do they really stabilize mast cells?

  Alessandro Lambiase, Alessandra Micera, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: Multiple action drugs, such as azelastine, epinastine, ketotifen and olopatadine, have recently been suggested to combine antihistaminic effect, mast cell stabilization and anti-inflammatory action. This pharmaceutical class is, therefore, rapidly becoming the first choice for prevention and treatment for allergic conjunctivitis. Increasing in-vitro studies have been performed to investigate the mast-cell-stabilizing effect of multiple action drugs. Most of the study results agree that these drugs are able to inhibit histamine and several neoformed mediators, including cytokines and arachidonic acid-derived products, from mast cells. However, the mechanisms of action have not yet fully been elucidated. Most of the results from clinical trials as well as the in-vivo experimental studies, including the conjunctival provocation model, support the evidence of a stabilizing effect of these drugs. Evidence of a different inhibitory effect of multiple action compounds on the pro-inflammatory mediators released from the mast cells suggests the possibility to target different phases of the allergic reaction, leading to a potential improvement in the management of allergic patients.
  Current Opinion in Allergy and Clinical Immunology 08/2009; 9(5):454-65. · 4.11 Impact Factor
• Article: In vitro evidence of nerve growth factor effects on human conjunctival epithelial cell differentiation and mucin gene expression.

  Alessandro Lambiase, Alessandra Micera, Graziella Pellegrini, Daniela Merlo, Paolo Rama, Michele De Luca, Sergio Bonini, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: Mucins released into the tear film are crucial to maintaining a healthy ocular surface. Alterations in goblet cell numbers and mucin secretion are observed in chronic ocular surface inflammatory diseases. Nerve growth factor (NGF) plays a crucial role in healing and inflammation of the ocular surface. The aim of this study was to evaluate in vitro the effect of NGF on conjunctival goblet cell differentiation and mucin production and secretion. Human conjunctival epithelial cells were exposed to increasing NGF concentrations (1 to 250 ng/mL) and analyzed to quantify cell growth (MTT/Ki67/BrdU), goblet cell differentiation (PAS/MUC5AC confocal staining), and mucin mRNA expression (real-time PCR). Secreted and cellular MUC5AC were also analyzed by sandwich-ELISA and FACS, respectively. To confirm the biological effects of NGF, the same evaluations were performed on primary cultures, and changes in markers of stemness (p63) and commitment (14-3-3 sigma) were also investigated. In cell cultures, NGF induced a dose-dependent increase of goblet cell numbers, MUC5AC production, storage, and release. Additionally, in primary cultures, NGF induced an increase of abortive colonies and 14-3-3 sigma protein, and a decrease of p63 mRNA and protein, suggesting a differentiating effect of NGF on human conjunctival epithelium. These findings show that NGF might play a role in the complex mechanism leading to conjunctival epithelium differentiation and mucin secretion. In addition to the known roles of NGF in promoting ocular surface healing and sensitivity, its effects on conjunctival goblet cells support a rationale to investigate the therapeutic effectiveness of NGF in dry eye disease.
  Investigative ophthalmology & visual science 05/2009; 50(10):4622-30. · 3.43 Impact Factor
• Source

  Available from: Sergio Bonini

  Article: Nerve growth factor modulates toll-like receptor (TLR) 4 and 9 expression in cultured primary VKC conjunctival epithelial cells.

  Alessandra Micera, Barbara Stampachiacchiere, Eduardo Maria Normando, Alessandro Lambiase, Sergio Bonini, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: To investigate if nerve growth factor (NGF) might modulate toll-like receptor (TLR) 4 and 9 expression in primary cultures of vernal keratoconjunctivitis (VKC)-derived conjunctival epithelial cells (VKC-ECs). Primary cultures of ECs were established from VKC (n=7) and healthy-control (n=5) conjunctival specimens. Primary untouched and short-term NGF-exposed VKC-ECs were analyzed for molecular (relative real-time PCR) and biochemical (confocal and fluorescence-activated cell sorting analysis of TLR4 and TLR9 expression. Data were compared to their untreated as well as stimulated healthy-control counterparts. Conditioned media were analyzed for interferon (IFN)-gamma, interleukin (IL)-4, IL-10, and IL-12 p40 secretion. Primary untouched VKC-ECs showed TLR4 increase and TLR9 decrease compared to their healthy-control counterparts. NGF exposure resulted in a strong upregulation of TLR4 and a moderate upregulation of TLR9 in few passages VKC-ECs. Both TLR4 and TLR9 upregulation occurred in a dose-dependent fashion and were supported by biochemical analysis. NGF triggered a dose-response release of IL-10 in VKC-ECs conditioned media, an effect not detected for IL-4, IL-12 p40, and IFN-gamma. Our data indicate that NGF is able to induce TLR4/TLR9 overexpression in VKC-ECs. These cells exhibited poor IL-4, IL-12 p40, and IFN-gamma responses to NGF, while a significant IL-10 decreased secretion was detected. The different NGF-induced TLR response between VKC and healthy-control conjunctival ECs as well as the different cytokine response might reflect a different pattern of cell activation according to the state of VKC.
  Molecular vision 01/2009; 15:2037-44. · 2.20 Impact Factor
• Source

  Available from: PubMed Central

  Article: T-helper 17 lymphocytes in ocular cicatricial pemphigoid.

  Alessandro Lambiase, Alessandra Micera, Flavio Mantelli, Caterina Moretti, Antonio Di Zazzo, Eleonora Perrella, Sergio Bonini, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: T-helper 17 lymphocytes (Th17) were identified in the healthy conjunctiva and in patients with ocular cicatricial pemphigoid (OCP), a disease characterized by chronic ocular surface inflammation. Conjunctival biopsies and blood samples were obtained from 10 patients with OCP (4 males, 6 females; 57-90 years of age) and 6 age/sex matched healthy subjects. Conjunctival samples were immunostained with anti-human IL17/CD4 antibodies and stained cells were then counted by confocal microscopy in three 60X field images per each sample. Mononuclear cells were isolated from both OCP and healthy blood samples and evaluated for IL17 and CD4 by FACS. IL17, TGF-beta, IL4, and IFN-gamma levels were determined in plasma of OCP and healthy patients by ELISA. The presence of Th17 lymphocytes in conjunctival biopsies was significantly (p<0.05) increased in patients with OCP (14.9+/-12.8 cells per microscopic field) compared to healthy subjects (0.5+/-0.8 cells per microscopic field). Th17 lymphocytes comprised 72% of CD4(+) cells in four stage-III OCP conjunctival samples. No significant difference was observed for IL17 in peripheral blood of OCP versus healthy subjects. In this study, we report an increased localization of Th17 lymphocytes in OCP conjunctiva, not accompanied by similar findings in peripheral blood. This finding suggests an increased recruitment of Th17 lymphocytes in conjunctiva and/or a dysfunctional local immune response in the chronically inflamed conjunctiva of OCP. Our findings are in line with previously reported evidence demonstrating that Th17 cells play a critical pathogenic role in mucosal autoimmunity.
  Molecular vision 01/2009; 15:1449-55. · 2.20 Impact Factor
• Article: The role of neuromediators in ocular allergy.

  Alessandra Micera, Alessandro Lambiase, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: To describe and highlight the critical role of neuromediators in ocular allergy, in view of the recent findings indicating that particularly neuropeptides and neurotrophins exert a pivotal role in the complex network of neurogenic inflammation occurring in ocular allergy. Neuropeptides and neurotrophins represent an interesting group of neuromediators actively taking place in allergic response. Mainly released at the neuronal edge or from immune and/or structural cells, neuropeptides and neurotrophins might either upregulate or downregulate inflammatory processes. These mediators contribute to ocular allergy by acting directly on cells responsible for both early and late phase reactions as well as on epithelial cells and fibroblasts, leading to either self-limiting or chronicity states. This short review surveys our current knowledge on the functional activity showed by substance P, calcitonin gene-related peptide, vasoactive intestinal peptide, neuropeptide Y and nerve growth factor and integrates their opposing effects in the context of neuroimmune interactions during ocular allergy. In addition, the review will provide implications for the clinical situation and/or future strategies for treating the most severe chronic forms of ocular allergy.
  Current Opinion in Allergy and Clinical Immunology 11/2008; 8(5):466-71. · 4.11 Impact Factor
• Article: Retinal p75 and bax overexpression is associated with retinal ganglion cells apoptosis in a rat model of glaucoma.

  Marco Coassin, Alessandro Lambiase, Valentina Sposato, Alessandra Micera, Stefano Bonini, Luigi Aloe
  [show abstract] [hide abstract]
  ABSTRACT: The administration of neurotrophins has been clearly demonstrated to support survival of retina cells during a variety of insults. Increased levels of neurotrophins, such as the nerve growth factor (NGF), have been found in experimental models of glaucoma. Nevertheless, loss of retinal cells does occur in the course of ocular hypertension. Therefore, this study sought to address whether timely changes in NGF and its receptors, trkA(NGFR) and p75(NTR), might explain the progression of retinal damage during experimental glaucoma. A well-characterized technique to induce glaucoma in rats was utilized. The animals were sacrificed after 10, 20 and 35 days from induction of glaucoma. Retinal ganglion cell (RGC) apoptosis, retinal expression of NGF protein as well as Bcl-2, Bax, trkA(NGFR) and p75(NTR) transcript expression were detected. The balance between trkA(NGFR) and p75(NTR) was examined, considering their anti- and pro-apoptotic role in cell death, respectively. We demonstrated that in our model of experimental glaucoma, the loss of retinal ganglion cells (RGCs) is accompanied by a timely increase of retinal NGF. Moreover, we found that the trkA(NGFR)/p75(NTR) mRNA ratio and the Bcl-2/Bax mRNA were both decreased, indicating a p75(NGFR) and Bax over-expression. Retinal NGF is over-expressed in experimental glaucoma, but this NGF increase is not sufficient to support survival of RGCs. The failure of NGF trophic support might be associated with the progressive up-regulation of p75(NTR) in relation to trkA(NGFR).
  Albrecht von Graæes Archiv für Ophthalmologie 09/2008; 246(12):1743-9. · 2.17 Impact Factor
• Article: Human idiopathic epiretinal membranes express NGF and NGF receptors.

  Simona Minchiotti, Barbara Stampachiacchiere, Alessandra Micera, Alessandro Lambiase, Guido Ripandelli, Bernardo Billi, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: Glial cells and fibroblasts (FBs) play a key role in epiretinal membrane (ERM) development and progression. Myofibroblasts (myoFBs), arising from these cells, can lead to the hypertrophic scars and tissue contraction observed in ERMs. Nerve growth factor (NGF) and transforming growth factor-beta1 (TGF-beta1) play a crucial role in FB activities. Therefore, the authors evaluated myoFBs in ERMs and NGF, trkA(NGFR and p75(NTR) expression, as well as TGF-beta1/TGF-betaRII levels in both ERMs and vitreous. Eight idiopathic ERMs and vitreous were obtained from patients at the time of vitrectomy for macular pucker. Ten control vitreous were from donors. Biochemical and molecular analyses were performed to identify alpha-smooth muscle actin (alpha-SMA, a defined myoFB marker), NGF, trkA(NGFR)/p75(NTR), and TGF-beta1/TGF-betaRII. Every idiopathic ERM displayed alpha-SMA positive myoFBs, expressing NGF, trkA(NGFR), and p75(NTR). ERM vitreous showed a significant decrease in NGF protein coupled with a TGF-beta1 increase. In addition, vitreous cells showed an increase in trkA(NGFR)/p75(NTR) mRNA associated with a decrease in TGF-betaRII mRNA. Idiopathic ERMs were characterized by myoFBs. The expression of NGF, trkA, and p75 in local myoFBs associated with changes in ERM vitreous NGF suggests an involvement of NGF, as previously reported for TGF-beta1, in the evolution and myoFB-mediated contractile activity of ERMs.
  Retina 05/2008; 28(4):628-37. · 2.81 Impact Factor
• Article: Preliminary evidence of the efficacy of probiotic eye-drop treatment in patients with vernal keratoconjunctivitis.

  Alfonso Iovieno, Alessandro Lambiase, Marta Sacchetti, Barbara Stampachiacchiere, Alessandra Micera, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: Probiotics have been shown to improve allergic inflammation. The aim of this study was to evaluate the efficacy of Lactobacillus Acidophilus eye-drops in controlling signs and symptoms of vernal keratoconjunctivitis (VKC). Seven patients (mean age 11.8 +/- 4.3; five M, two F) with mild to moderate VKC were included in the study. Lactobacillus Acidophilus diluted in saline solution (2 x 10(8) CFU/ml) was administrated as eye-drops four times daily for 4 weeks in both eyes. Clinical signs (conjunctival hyperemia, chemosis, secretion, Trantas dots, superficial punctuate keratitis) and symptoms (itching, photophobia, burning, tearing) were evaluated and scored from 0 to 3 at baseline, after 2 and 4 weeks of treatment. Total sign (TSS) and symptom (TSyS) scores were calculated. Conjunctival impression cytology was performed in three patients at baseline and after 4 weeks of treatment, in order to evaluate the expression of ICAM-1 and TLR-4. In the six out of seven patients who completed the study, symptoms were significantly improved after both 2 weeks (TSyS: baseline 6.7 +/- 0.9 vs 4.1 +/- 1.2; p = 0.017) and 4 weeks (TSyS: baseline 6.7 +/- 0.9 vs 3.6 +/- 1.2, p = 0.011) of treatment. A significant improvement of clinical signs was observed after 4 weeks of treatment (TSS: baseline 7.5 +/- 1.6 vs 3.9 +/- 1.7, p = 0.034) but not after 2 weeks of treatment (TSS: baseline 7.5 +/- 1.6 vs 5.3 +/- 1.5; NS). In particular, photophobia was significantly reduced (2 +/- 0.6 vs 1 +/- 0.3; p = 0.023) at 2 weeks, while at 4 weeks the scores for itching (1.8 +/- 0.3 vs 1 +/- 0.3), tearing (1.6 +/- 0.4 vs 0.8 +/- 0.2), conjunctival hyperemia (2.3 +/- 0.2 vs 1.4 +/- 0.5) and chemosis (1.2 +/- 0.4 vs 0.4 +/- 0.4) were significantly lower compared to baseline. A down-regulation of ICAM-1 and TLR-4 was observed in two patients showing clinical improvement after 4 weeks of treatment. Our open pilot study showed that 1-month treatment with probiotic eye-drops improves signs and symptoms in patients with VKC. Additional double-blind controlled clinical trials with a larger sample of patients are needed to confirm the effects of topical Lactobacilli on VKC patients.
  Albrecht von Graæes Archiv für Ophthalmologie 04/2008; 246(3):435-41. · 2.17 Impact Factor
• Article: Topical treatment with nerve growth factor in an animal model of herpetic keratitis.

  Alessandro Lambiase, Marco Coassin, Nicola Costa, Paolo Lauretti, Alessandra Micera, Emiliano Ghinelli, Luigi Aloe, Paolo Rama, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: In vitro and in vivo studies demonstrated the antiviral efficacy of nerve growth factor (NGF) and its cyto-protective effect in herpes simplex virus (HSV)-infected cells. The aims of this study were to evaluate the role of endogenous NGF in HSV corneal infection, and the effects of topical NGF treatment on herpetic keratitis. Herpetic keratitis was induced in 40 rabbits with the HSV-1 McKrae strain. Animals were divided into four groups, and treated with topical neutralizing anti-NGF antibodies, NGF, acyclovir or balanced salt solution (BSS) respectively. The clinical course of HSV keratitis was evaluated and scored by slit-lamp examination. In addition, biochemical (immunohistochemistry for glycoprotein D) and molecular (nested PCR for glycoprotein D) analyses were carried out to estimate viral replication. Treatment with anti-NGF antibodies induced a more severe keratitis associated with increased biochemical and molecular markers of active viral replication. Two animals in this group developed lethal HSV encephalitis. Conversely, topical treatment with NGF induced a significant amelioration of clinical and laboratory parameters when compared to the BSS treated group (control). No significant differences were observed between NGF- and acyclovir-treated groups. This study demonstrated the crucial role of endogenous NGF in herpetic keratitis. The comparable effects of NGF and acyclovir confirm the antiviral activity of NGF, and indicate a potential use of topical NGF in herpetic keratitis.
  Albrecht von Graæes Archiv für Ophthalmologie 02/2008; 246(1):121-7. · 2.17 Impact Factor
• Article: MUC5AC overexpression in tear film of neonates.

  Flavio Mantelli, Eloisa Tiberi, Alessandra Micera, Alessandro Lambiase, Federica Visintini, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: Full-term neonates produce tears normally, but neonatal tear film is modified to resist evaporation with a thick lipid layer that allows lower spontaneous blink rates. This adaptation presumably prevents drying of the ocular surface during long inter-blink periods. However, tear-film stability is not only based on the integrity of the lipid layer, but also reflects properties of the underlying mucus layer. Characteristics of the neonatal mucus tear-film layer have not yet been described. Tear samples were obtained from eight full-term healthy neonates (four males, four females, mean age 1.7 +/- 0.5 days) and eight healthy adult controls (four males, four females, mean age 26.3 +/- 2.5 years). Characterization of tear samples' total proteins was obtained by spectrophotometry. Western blot for major secretory mucin MUC5AC was performed on the samples. Blink rate in the neonates and adults enrolled in the study was also observed and recorded. Using the same procedure, the amount of tears collected was significantly greater in adults than in neonates (p < .01). Western Blot performed on neonatal tear samples showed a significant 76.8% increase in the expression of major secretory mucin MUC5AC as compared to healthy adult controls (p < .001). Mean blink rate recorded in neonates was significantly lower than in adults (p < .001), with a mean 1.6 +/- 0.5 blinks per minute and a mean interblink time of 33 +/- 9 seconds. As far as we are aware this is the first description of the mucus tear-film layer in neonates. The greater tear-film stability in neonates has been so far attributed to a thicker lipid layer. In our study, we show that a concomitant increase in MUC5AC protein expression in tears is present and may contribute to this greater stability; therefore, both mucus and lipid layer should be considered while evaluating tear film stability in neonates.
  Albrecht von Graæes Archiv für Ophthalmologie 10/2007; 245(9):1377-81. · 2.17 Impact Factor
• Article: NGF topical application in patients with corneal ulcer does not generate circulating NGF antibodies.

  Alessandro Lambiase, Marco Coassin, Valentina Sposato, Alessandra Micera, Marta Sacchetti, Stefano Bonini, Luigi Aloe
  [show abstract] [hide abstract]
  ABSTRACT: Nerve growth factor (NGF) is the prototype of the neurotrophin family and promotes the survival of specific populations of neurons, stimulates their morphological differentiation and regulates neuronal gene expression. Since its discovery, NGF attracted clinicians for its potential application in the treatment of neurological disorders. Recent studies demonstrated murine NGF eye drops may be successfully used to treat neurotrophic keratopathy. Neurotrophic keratopathy is a degenerative corneal disease caused by an impairment of the trigeminal nerve, which leads to corneal epithelial defect, ulcer, and perforation. NGF topical application induced complete ulcer healing and a recovery of the corneal sensitivity. The intent of this study is to address the question if murine NGF topical treatment stimulates the production of circulating anti-NGF antibodies. We evaluated patients with neurotrophic keratopathy who underwent topical therapy with NGF eye drops for possible ocular or systemic adverse effects in the course of 16-72 months follow up. None of the patients suffered any adverse reaction, except for mild and transient conjunctival hyperaemia and photophobia. Moreover, none of the treated patients developed circulating antibodies to NGF.
  Pharmacological Research 08/2007; 56(1):65-9. · 4.44 Impact Factor
• Source

  Available from: Costanzo Moretti

  Article: Conjunctival mucin deficiency in complete androgen insensitivity syndrome (CAIS).

  Flavio Mantelli, Costanzo Moretti, Alessandra Micera, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: Sex steroid hormones are essential for a healthy ocular surface and the androgen receptor impairment found in patients with complete androgen insensitivity syndrome (CAIS) has been described to cause meibomian gland dysfunction and functional dry eye for lipid tear film layer instability. However, it has not been reported if the mucous layer is also affected. A 37-year-old CAIS patient with persistent symptoms of dry eye underwent ophthalmological examination and was evaluated for qualitative and quantitative tear function tests and conjunctival cytology. Samples obtained from the conjunctival epithelium were stained for histology and immunohistochemistry and compared with three age-matched female controls. Western blot and relative real-time RT-PCR for MUC1 and MUC5AC were also performed on these samples. Immunohistochemistry, Western blot and relative real-time RT-PCR showed a decrease in the expression of MUC1 and MUC5AC in CAIS. Changes in the tear film mucous layer were accompanied by a reduction in the tear film break up time test. This is the first report describing mucous layer alteration associated with androgen receptor impairment. Decreased mucin levels contribute in explaining the tear film instability in CAIS and should be considered an additional cause of dry eye in sex steroid hormone pathology.
  Albrecht von Graæes Archiv für Ophthalmologie 07/2007; 245(6):899-902. · 2.17 Impact Factor
• Source

  Available from: Sergio Bonini

  Article: Itchy-dry eye associated with polycystic ovary syndrome.

  Stefano Bonini, Flavio Mantelli, Costanzo Moretti, Alessandro Lambiase, Sergio Bonini, Alessandra Micera
  [show abstract] [hide abstract]
  ABSTRACT: The authors aimed to define the ocular symptomatology of women with polycystic ovaries and hyperandrogenism. Prospective, observational case series. Of the 62 consecutive patients with an ultrasonographic diagnosis of polycystic ovary (PCO), 16 were identified as having clinical and biochemical signs of hyperandrogenism. All women with a history of ocular symptoms (20/62 total patients [32.3%], 15/16 polycystic ovary syndrome (PCOS) patients [93.7%], and 5/46 PCO patients [10.8%]) underwent a complete eye examination with conjunctival impression cytologic sampling. Clinical measurements of tear function (tear film break-up time [BUT], Schirmer I test) were completed along with analysis of conjunctival goblet cell number, conjunctival immunostaining, and reverse-transcriptase polymerase chain reaction for the mucins MUC1 and MUC5AC. Clinical, histologic, and biochemical data of patients with PCOS were compared statistically with that of patients with PCO and with eight age- and gender-matched healthy controls. Eight of the most severely affected patients received systemic antiandrogen therapy and underwent further ocular evaluation four months after systemic therapy. Women with PCOS had greater conjunctival hyperemia (P < .001), dryness (P < .001), itching (P < .001), mucous discharge (P < .001), and contact lens intolerance (P < .001) than patients with PCO. Patients with PCOS had a significant reduction of the tear film BUT accompanied by a significant increase in goblet cell number and conjunctival MUC5AC messenger ribonucleic acid expression compared with both PCO patients and healthy subjects. Evaluation of the ocular surface should be considered in patients with PCO or PCOS. Women with PCOS were more likely to have itchy-dry eyes, decreased tear film BUT, and increased goblet cell density.
  American Journal of Ophthalmology 06/2007; 143(5):763-771. · 4.22 Impact Factor
• Source

  Available from: Sergio Bonini

  Article: Nerve growth factor has a modulatory role on human primary fibroblast cultures derived from vernal keratoconjunctivitis-affected conjunctiva.

  Alessandra Micera, Alessandro Lambiase, Barbara Stampachiacchiere, Roberto Sgrulletta, Eduardo Maria Normando, Sergio Bonini, Stefano Bonini
  [show abstract] [hide abstract]
  ABSTRACT: To evaluate the role of nerve growth factor (NGF) in remodeling processes of vernal keratoconjunctivitis (VKC). VKC is a chronic inflammatory disorder of the conjunctiva and is characterized by marked tissue remodeling. NGF, a pleiotrophic factor with documented profibrogenic activities, is produced by inflammatory and structural cells populating the VKC conjunctiva and is increased in the serum and tears of VKC patients. Primary cultures of VKC-derived fibroblasts (VKC-FBs) were exposed to increasing NGF concentrations (1-500 ng/ml) to evaluate and compare the expression of alpha-smooth muscle actin (alphaSMA, a defining myofibroblast marker), collagens (types I and IV), and metalloproteinases and tissue inhibitors (MMP9/TIMP1, MMP2/TIMP2) at the biochemical as well as molecular levels. Endogenous NGF was increased in the VKC-FB supernatant, as compared to healthy-FB supernatant. VKC-FBs expressed alphaSMA and increased types I and IV collagens. VKC-FBs, and in particular all alphaSMA positive cells, expressed both trkA(NGFR) and p75(NTR), while healthy-FBs only expressed trkA(NGFR). Exogenous NGF did not change alphaSMA expression, while alphaSMA expression was enhanced by specific neutralization of p75(NTR). NGF (10 ng/ml) exposure significantly decreased type I collagen expression, without affecting type IV collagen, and increased MMP9mRNA and protein. The autocrine modulation of differentiation and response of VKC-FBs to NGF exposure with downregulation of type I collagen and upregulation of MMP9 expression supports a relevant role for NGF in tissue remodeling of VKC.
  Molecular vision 02/2007; 13:981-7. · 2.20 Impact Factor