Annalisa Marcuzzi - Academia.edu (original) (raw)

Papers by Annalisa Marcuzzi

Molecular Medicine Reports, 2016

Mevalonate kinase deficiency (MKD) is an autosomal recessive auto-inflammatory disease, caused by... more Mevalonate kinase deficiency (MKD) is an autosomal recessive auto-inflammatory disease, caused by impairment of the mevalonate pathway. Although the molecular mechanism remains to be elucidated, there is clinical evidence suggesting that other regulatory genes may be involved in determining the phenotype. The identification of novel target genes may explain non-homogeneous genotype-phenotype correlations, and provide evidence in support of the hypothesis that novel regulatory genes predispose or amplify deregulation of the mevalonate pathway in this orphan disease. In the present study, DNA samples were obtained from five patients with MKD, which were then analyzed using whole exome sequencing. A missense variation in the PEX11γ gene was observed in homozygosis in P2, possibly correlating with visual blurring. The UNG rare gene variant was detected in homozygosis in P5, without correlating with a specific clinical phenotype. A number of other variants were found in the five analyzed DNA samples from the MKD patients, however no correlation with the phenotype was established. The results of the presents study suggested that further analysis, using next generation sequencing approaches, is required on a larger sample size of patients with MKD, who share the same MVK mutations and exhibit 'extreme' clinical phenotypes. As MVK mutations may be associated with MKD, the identification of specific modifier genes may assist in providing an earlier diagnosis.

Molecular Medicine Reports, 2015

Aminobisphosphonate aledronate is a compound commonly used clinically for the treatment of osteop... more Aminobisphosphonate aledronate is a compound commonly used clinically for the treatment of osteoporosis and other bone diseases, as a result of it preventing bone resorption. However, in previous years it has also been used to obtain cellular and animal models of a rare genetic disorder termed Mevalonate Kinase Deficiency (MKD). MKD is caused by mutations affecting the mevalonate kinase enzyme, in the cholesterol pathway and alendronate can be used to biochemically mimic the genetic defect as it inhibits farnesyl pyrophosphate synthase in the same pathway. Despite evidence in favor of the inhibition exerted on the mevalonate pathway, there is at least one clinical case of MKD in which alendronate improved not only skeletal and bone fractures, as expected, but also MKD clinical features. Based on this finding, the present study assessed the anti‑inflammatory properties of this aminobisphosphonate in vitro. No anti‑inflammatory effects of alendronate were observed in the in vitro experiments. Since MKD lacks specific treatments, these results may assist scientists and physicians in making the decision as to the most suitable choice of therapeutic compounds for this neglected disease.

Apoptosis, 2015

Mevalonate kinase deficiency is a rare disease whose worst manifestation, characterised by severe... more Mevalonate kinase deficiency is a rare disease whose worst manifestation, characterised by severe neurologic impairment, is called mevalonic aciduria. The progressive neuronal loss associated to cell death can be studied in vitro with a simplified model based on a biochemical block of the mevalonate pathway and a subsequent inflammatory trigger. The aim of this study was to evaluate the effect of the mevalonate blocking on glial cells (BV-2) and the following effects on neuronal cells (SH-SY5Y) when the two populations were cultured together. To better understand the cross-talk between glial and neuronal cells, as it happens in vivo, BV-2 and SH-SY5Y were co-cultured in different experimental settings (alone, transwell, direct contact); the effect of mevalonate pathway biochemical block by Lovastatin, followed by LPS inflammatory trigger, were evaluated by analysing programmed cell death and mitochondrial membrane potential, cytokines' release and cells' morphology modifications. In this experimental condition, glial cells underwent an evident activation, confirmed by elevated pro-inflammatory cytokines release, typical of these disorders, and a modification in morphology. Moreover, the activation induced an increase in apoptosis. When glial cells were co-cultured with neurons, their activation caused an increase of programmed cell death also in neuronal cells, but only if the two populations were cultured in direct contact. Our findings, being aware of the limitations related to the cell models used, represent a preliminary step towards understanding the pathological and neuroinflammatory mechanisms occurring in mevalonate kinase diseases. Contact co-culture between neuronal and microglial cells seems to be a good model to study mevalonic aciduria in vitro, and to contribute to the identification of potential drugs able to block microglial activation for this orphan disease. In fact, in such a pathological condition, we demonstrated that microglial cells are activated and contribute to neuronal cell death. We can thus hypothesise that the use of microglial activation blockers could prevent this additional neuronal death.

Clinical Immunology, 2015

There is a lack of reliable animal models for the study of the rare auto-inflammatory disease mev... more There is a lack of reliable animal models for the study of the rare auto-inflammatory disease mevalonate kinase deficiency (MKD). The one most frequently used is a biochemical model, obtained by treating BALB/c mice in order to block the mevalonate pathway, thus attempting to reproduce the inflammatory pattern presented in patients. This study aims to assess the role played in pathology by the inflammasome and the reliability of this model. We mimicked MKD using two different mice strains (BALB/c and C57BL/6), evaluating typical inflammatory markers of MKD and inflammasome modulation. Without significant differences, both strains exhibited a general MKD-like inflammation, including the modulation of the molecular platform inflammasome, mimicking the characteristics observed in human patients. Although with some limitations, the mouse model appears robust and suitable for studying MKD. Results do not seem to vary with the mouse strain used, and appear to be treatment-dependent. Finally, in vivo inflammasome activation was assessed for the first time here.

International Journal of Developmental Neuroscience, 2012

Mevalonic aciduria (MA), the most severe form of mevalonate kinase deficiency (MKD), is still an ... more Mevalonic aciduria (MA), the most severe form of mevalonate kinase deficiency (MKD), is still an orphan drug disease and the pathogenetic mechanisms underlying neuronal dysfunction is still poorly understood. In our study we have investigated the apoptotic mechanism mediated by the exposure of the cultured neuroblastoma cell line, SH-SY5Y, to lovastatin in absence or in presence of the isoprenoid, geranylgeraniol, with the aim of unraveling the pathogenesis of MA. Lovastatin, blocks the mevalonate pathway inhibiting the 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CR), an enzyme of the mevalonate pathway upstream the mevalonate kinase enzyme, reproducing biochemical features similar to those found in MKD.

Journal of Cardiovascular Translational Research, 2013

Since contradictory findings have been reported on potential effects of statins in modulating the... more Since contradictory findings have been reported on potential effects of statins in modulating the inflammatory response, we have analysed the biological activity of lovastatin both in vitro using the Raw 264.7 murine macrophagic cell line and in vivo using BALB/c mice. When added to Raw 264.7 cells in combination with lipopolysaccharide, lovastatin significantly potentiated the release of interleukin-1β, interleukin-6 and interleukin-12 with respect to lipopolysaccharide alone and showed an additive effect on the release of nitric oxide. Similarly, when lovastatin was intraperitoneally administrated to BALB/c mice, it did not induce any proinflammatory effect when used alone, but it significantly potentiated the pro-inflammatory activity of lipopolysaccharide, in terms of number of intraperitoneal cells and serum levels of serum amyloid A, interleukin-1β, interleukin-6 and interleukin-12. A potential clinical implication of our study is that lovastatin might exert a pro-inflammatory activity in subjects affected by inflammatory processes, with clinically evident or subclinical infections.

BioMed Research International, 2013

Mevalonate Kinase Deficiency (MKD) is a rare autosomal recessive inborn disorder of cholesterol b... more Mevalonate Kinase Deficiency (MKD) is a rare autosomal recessive inborn disorder of cholesterol biosynthesis caused by mutations in the mevalonate kinase (MK) gene, leading to MK enzyme decreased activity. The consequent shortage of mevalonate-derived isoprenoid compounds results in an inflammatory phenotype, caused by the activation of the NALP3 inflammasome that determines an increased caspase-1 activation and IL-1 release. In MKD, febrile temperature can further decrease the residual MK activity, leading to mevalonate pathway modulation and to possible disease worsening. We previously demonstrated that the administration of exogenous isoprenoids such as geraniol or the modulation of the enzymatic pathway with drugs, such as Tipifarnib, partially rescues the inflammatory phenotype associated with the defective mevalonic pathway. However, it has not been investigated yet how temperature can affect the success of these treatments. Thus, we investigated the effect of temperature on primary human monocytes from MKD patients. Furthermore the ability of geraniol and Tipifarnib to reduce the abnormal inflammatory response, already described at physiological temperature in MKD, was studied in a febrile condition. We evidenced the role of temperature in the modulation of the inflammatory events and suggested strongly considering this variable in future researches aimed at finding a treatment for MKD.

Pediatric Research, 2008

Mevalonate kinase deficiency (MKD) is a rare disorder characterized by recurrent inflammatory epi... more Mevalonate kinase deficiency (MKD) is a rare disorder characterized by recurrent inflammatory episodes and, in most severe cases, by psychomotor delay. Defective synthesis of isoprenoids has been associated with the inflammatory phenotype in these patients, but the molecular mechanisms involved are still poorly understood, and, so far, no specific therapy is available for this disorder. Drugs like aminobisphosphonates, which inhibit the mevalonate pathway causing a relative defect in isoprenoids synthesis, have been also associated to an inflammatory phenotype. Recent data asserted that cell inflammation could be reversed by the addition of some isoprenoids, such as geranylgeraniol and farnesyl pyrophosphate. In this study, a mouse model for typical MKD inflammatory episode was obtained treating BALB/c mice with aminobisphosphonate alendronate and bacterial muramyldipeptide. The effect of exogenous isoprenoids-geraniol, farnesol, and geranylgeraniol-was therefore evaluated in this model. All these compounds were effective in preventing the inflammation induced by alendronate-muramyldipeptide, suggesting a possible role for these compounds in the treatment of MKD in humans.

Pediatric Research, 2013

Mevalonate kinase deficiency (MKD) is a rare genetic autoinflammatory disease caused by blocking ... more Mevalonate kinase deficiency (MKD) is a rare genetic autoinflammatory disease caused by blocking of the enzyme mevalonate kinase in the pathway of cholesterol and isoprenoids. The pathogenic mechanism originating an immune response in MKD patients has not been clearly understood. We investigated the dysregulation of expression of selected cytokines and chemokines in the serum of MKD patients. The results have been compared with those observed in an MKD mouse model obtained by treating the mice with aminobisphosphonate, a molecule that is able to inhibit the cholesterol pathway, mimicking the genetic block characteristic of the disease. Interleukin (IL)-1β, IL-5, IL-6, IL-9, IL-17, granulocyte colony-stimulating factor, monocyte chemotactic protein-1, tumor necrosis factor-α, and IL-4 expression were dysregulated in sera from MKD patients and mice. Moreover, geraniol, an exogenous isoprenoid, when administered to MKD mice, restored cytokines and chemokines levels with values similar to those of untreated mice. Our findings, which were obtained in patients and a mouse model mimicking the human disease, suggest that these cytokines and chemokines could be MKD specific and that isoprenoids could be considered as potential therapeutic molecules. The mouse model, even if with some limitations, was robust and suitable for routine testing of potential MKD drugs.

Medical Hypotheses, 2012

Crohn&amp... more Crohn's disease (CD) is a multifactorial disease, in which environmental, microbial and genetic factors play important roles. CD is characterized by a chronic granulomatous inflammation by necrotic scarring with aspects of full-thickness wall. In spite of affecting mainly young adults, sometimes, CD can be present in the first year of life (early onset Crohn disease, EOCD) showing an unpredictable course and being often more severe than at older ages. In this paper we propose the hypothesis that EOCD patients should be analyzed using a Mendelian approach with family studies aimed to identify new loci directly involved in the early onset Crohn's disease. So we will leave the classic association study approach used until now for the identification of genes responsible for susceptibility to CD and propose linkage family analysis as alternative and powerful tool for the identification of new genetic variants associated with familiar cases of EOCD.

Medical Hypotheses, 2013

Mevalonate Kinase Deficiency (MKD) is an autosomal-recessively inherited disorder of cholesterol ... more Mevalonate Kinase Deficiency (MKD) is an autosomal-recessively inherited disorder of cholesterol biosynthesis with higher prevalence in the Netherlands and other North European countries. MKD is due to mutations in the second enzyme of mevalonate pathway (mevalonate kinase, MK/MVK) which results in reduced enzymatic activity and in the consequent shortage of downstream compounds. In most severe cases the deregulation of mevalonate pathway is associated with a decrease in serum cholesterol. More than 100 pathological mutations have been described in the MVK gene so far, and a founder effect has been hypothesized as responsible for the diffusion of the most frequent disease-associated mutations. In the acute phase of disease, patients affected with MKD present low cholesterol levels comparable to their basal physiologic conditions, already characterized by lower cholesterol levels when compared to healthy individuals. Low cholesterol levels are widely known to correlate with the reduction of cardiovascular events. We hypothesize a selective advantage for heterozygote carriers of the most frequent MVK mutations in those countries where the diet is characterized by high consumption of saturated animal fats rich in cholesterol. This could explain the maintenance in North European population of the main mutations leading to MKD and the distribution world-wide of these mutations that followed the migrations of North European populations.

International Immunopharmacology, 2010

The inhibition of mevalonate pathway through genetic defects (mevalonate kinase deficiency, MKD) ... more The inhibition of mevalonate pathway through genetic defects (mevalonate kinase deficiency, MKD) or pharmacologic drugs (aminobisphosphonates) causes a shortage of intermediate compounds and, in particular, of geranylgeranyl-pyrophosphate (GGPP) associated to the activation of caspase-1 and IL-1β release. Geraniol (GOH), farnesol (FOH), geranylgeraniol (GGOH) and menthol (MOH), due to their isoprenoid structure, are supposed to enter the mevalonate pathway and to by-pass the biochemical block, reconstituting the pathway. Considering the already known side effects of aminobisphosphonates, and the lack of a specific treatment for MKD, we evaluated the impact of these natural isoprenoids compounds in a RAW cell lines chemically treated with the aminobisphosphonate alendronate, and in monocytes isolated from 2 patients affected by MKD. GOH, FOH, GGOH and MOH were all capable to diminish inflammatory marker levels induced by LPS. These natural isoprenoids could be proposed as novel therapeutic approach for the still orphan drug MKD, but also considered for the evaluation of possible inflammatory side effects of aminobisphosphonates.

Inflammation Research, 2010

Objective The aim of this study was to evaluate, in a mouse model of mevalonate kinase deficiency... more Objective The aim of this study was to evaluate, in a mouse model of mevalonate kinase deficiency (MKD), the possible link between inflammatory symptoms and serum cholesterol levels. Materials and methods Balb/c mice were treated with alendronate and bacterial muramyl dipeptide. Body temperature, interleukin-1b (IL-1b) secretion and serum cholesterol levels were measured. Results An increased production of the pro-inflammatory cytokine IL-1b (p \ 0.05) and a rise in body temperature (p \ 0.05) was observed, while, in parallel, serum cholesterol concentration significantly decreased (p \ 0.05). These effects were completely reversed when animals were treated with exogenous isoprenoids. Conclusions In the mouse model of MKD, the inflammatory response is associated with a reduction in cholesterol levels, and hence this parameter could be used as an indicator of isoprenoid consumption. In addition, plant derived isoprenoids could represent candidate treatments for this disease.

Inflammation Research, 2012

A potential anti-inflammatory effect of clodronate--an aminobisphosphonate--was described to anta... more A potential anti-inflammatory effect of clodronate--an aminobisphosphonate--was described to antagonize the pro-inflammatory effects of the block in the mevalonate pathway, the main feature of a rare auto-inflammatory disease called mevalonate kinase deficiency (MKD). In this study we evaluated the potential anti-inflammatory effect of clodronate in MKD--a still orphan drug pediatric disease. We studied some biological parameters, nitric oxide production using Griess reagents and programmed cell death by flow cytometry, as common inflammatory parameters in MKD, in the presence of different doses of clodronate (1, 10 and 100 μM). In our cellular model and in monocytes from patients with MKD, clodronate induced an increase in programed cell death and nitric oxide production in comparison with non-treated cells. Our findings suggest that clodronate does not have an anti-inflammatory effect as previously reported but that it increases the epiphenomena of this pediatric disease.

Genomics, 2013

The knowledge of the human genome is in continuous progression: a large number of databases have ... more The knowledge of the human genome is in continuous progression: a large number of databases have been developed to make meaningful connections among worldwide scientific discoveries. This paper reviews bioinformatics resources and database tools specialized in disseminating information regarding genetic disorders. The databases described are useful for managing sample sequences, gene expression and post-transcriptional regulation. In relation to data sets available from genome-wide association studies, we describe databases that could be the starting point for developing studies in the field of complex diseases, particularly those in which the causal genes are difficult to identify.

Gene, 2012

Crohn disease (CD) is a multifactorial disorder affecting mainly young adults. Sometimes, however... more Crohn disease (CD) is a multifactorial disorder affecting mainly young adults. Sometimes, however, it can present in the first year of life (Early onset Crohn disease (EOCD)) showing an unpredictable course and can often be more severe than at older ages. Some cases have been associated to an underlying primary immunodeficiency such as IL10R deficiency. We studied the functional response to IL-10 and the genotype of IL-10 receptor in four patients with early onset crohn-like colitis. We found an IL10R variant, which may be associated with a decreased response to the cytokine in one patient. Further studies to determine its pathogenic effect should be performed. In addition IL-10 mediated inhibition of LPS-induced TNFα expression was measured in patient's monocytes.

Apoptosis, 2011

The inhibition of mevalonate pathway by the aminobisphosphonate alendronate (ALD) has been previo... more The inhibition of mevalonate pathway by the aminobisphosphonate alendronate (ALD) has been previously associated with an augmented lipopolysaccharideinduced interleukin-1beta (IL-1b) secretion in monocytes, as demonstrated in an auto-inflammatory disease known as mevalonate kinase deficiency (MKD). In this study we investigated the effect of ALD ? LPS on monocyte cell line (Raw 264.7) death. ALD strongly augmented LPSinduced programmed cell death (PCD) as well as IL-1b secretion in Raw murine monocytes, whereas necrosis was rather unaffected. ALD ? LPS induced caspase-3 activation. Inhibition of IL-1b stimulation partially restored cell viability. These findings suggest that the inhibition of mevalonate pathway, together with a bacterial stimulus, induce a PCD partly sustained by the caspase-3-related apoptosis and partly by caspase-1-associated pyroptosis. The involvement of pyroptosis is a novel hit in our cell model and opens discussions about its role in inflammatory cells with chemical or genetic inhibition of mevalonate pathway.

Molecular Medicine Reports, 2016

Mevalonate kinase deficiency (MKD) is an autosomal recessive auto-inflammatory disease, caused by... more Mevalonate kinase deficiency (MKD) is an autosomal recessive auto-inflammatory disease, caused by impairment of the mevalonate pathway. Although the molecular mechanism remains to be elucidated, there is clinical evidence suggesting that other regulatory genes may be involved in determining the phenotype. The identification of novel target genes may explain non-homogeneous genotype-phenotype correlations, and provide evidence in support of the hypothesis that novel regulatory genes predispose or amplify deregulation of the mevalonate pathway in this orphan disease. In the present study, DNA samples were obtained from five patients with MKD, which were then analyzed using whole exome sequencing. A missense variation in the PEX11γ gene was observed in homozygosis in P2, possibly correlating with visual blurring. The UNG rare gene variant was detected in homozygosis in P5, without correlating with a specific clinical phenotype. A number of other variants were found in the five analyzed DNA samples from the MKD patients, however no correlation with the phenotype was established. The results of the presents study suggested that further analysis, using next generation sequencing approaches, is required on a larger sample size of patients with MKD, who share the same MVK mutations and exhibit 'extreme' clinical phenotypes. As MVK mutations may be associated with MKD, the identification of specific modifier genes may assist in providing an earlier diagnosis.

Molecular Medicine Reports, 2015

Aminobisphosphonate aledronate is a compound commonly used clinically for the treatment of osteop... more Aminobisphosphonate aledronate is a compound commonly used clinically for the treatment of osteoporosis and other bone diseases, as a result of it preventing bone resorption. However, in previous years it has also been used to obtain cellular and animal models of a rare genetic disorder termed Mevalonate Kinase Deficiency (MKD). MKD is caused by mutations affecting the mevalonate kinase enzyme, in the cholesterol pathway and alendronate can be used to biochemically mimic the genetic defect as it inhibits farnesyl pyrophosphate synthase in the same pathway. Despite evidence in favor of the inhibition exerted on the mevalonate pathway, there is at least one clinical case of MKD in which alendronate improved not only skeletal and bone fractures, as expected, but also MKD clinical features. Based on this finding, the present study assessed the anti‑inflammatory properties of this aminobisphosphonate in vitro. No anti‑inflammatory effects of alendronate were observed in the in vitro experiments. Since MKD lacks specific treatments, these results may assist scientists and physicians in making the decision as to the most suitable choice of therapeutic compounds for this neglected disease.

Apoptosis, 2015

Mevalonate kinase deficiency is a rare disease whose worst manifestation, characterised by severe... more Mevalonate kinase deficiency is a rare disease whose worst manifestation, characterised by severe neurologic impairment, is called mevalonic aciduria. The progressive neuronal loss associated to cell death can be studied in vitro with a simplified model based on a biochemical block of the mevalonate pathway and a subsequent inflammatory trigger. The aim of this study was to evaluate the effect of the mevalonate blocking on glial cells (BV-2) and the following effects on neuronal cells (SH-SY5Y) when the two populations were cultured together. To better understand the cross-talk between glial and neuronal cells, as it happens in vivo, BV-2 and SH-SY5Y were co-cultured in different experimental settings (alone, transwell, direct contact); the effect of mevalonate pathway biochemical block by Lovastatin, followed by LPS inflammatory trigger, were evaluated by analysing programmed cell death and mitochondrial membrane potential, cytokines' release and cells' morphology modifications. In this experimental condition, glial cells underwent an evident activation, confirmed by elevated pro-inflammatory cytokines release, typical of these disorders, and a modification in morphology. Moreover, the activation induced an increase in apoptosis. When glial cells were co-cultured with neurons, their activation caused an increase of programmed cell death also in neuronal cells, but only if the two populations were cultured in direct contact. Our findings, being aware of the limitations related to the cell models used, represent a preliminary step towards understanding the pathological and neuroinflammatory mechanisms occurring in mevalonate kinase diseases. Contact co-culture between neuronal and microglial cells seems to be a good model to study mevalonic aciduria in vitro, and to contribute to the identification of potential drugs able to block microglial activation for this orphan disease. In fact, in such a pathological condition, we demonstrated that microglial cells are activated and contribute to neuronal cell death. We can thus hypothesise that the use of microglial activation blockers could prevent this additional neuronal death.

Clinical Immunology, 2015

There is a lack of reliable animal models for the study of the rare auto-inflammatory disease mev... more There is a lack of reliable animal models for the study of the rare auto-inflammatory disease mevalonate kinase deficiency (MKD). The one most frequently used is a biochemical model, obtained by treating BALB/c mice in order to block the mevalonate pathway, thus attempting to reproduce the inflammatory pattern presented in patients. This study aims to assess the role played in pathology by the inflammasome and the reliability of this model. We mimicked MKD using two different mice strains (BALB/c and C57BL/6), evaluating typical inflammatory markers of MKD and inflammasome modulation. Without significant differences, both strains exhibited a general MKD-like inflammation, including the modulation of the molecular platform inflammasome, mimicking the characteristics observed in human patients. Although with some limitations, the mouse model appears robust and suitable for studying MKD. Results do not seem to vary with the mouse strain used, and appear to be treatment-dependent. Finally, in vivo inflammasome activation was assessed for the first time here.

International Journal of Developmental Neuroscience, 2012

Mevalonic aciduria (MA), the most severe form of mevalonate kinase deficiency (MKD), is still an ... more Mevalonic aciduria (MA), the most severe form of mevalonate kinase deficiency (MKD), is still an orphan drug disease and the pathogenetic mechanisms underlying neuronal dysfunction is still poorly understood. In our study we have investigated the apoptotic mechanism mediated by the exposure of the cultured neuroblastoma cell line, SH-SY5Y, to lovastatin in absence or in presence of the isoprenoid, geranylgeraniol, with the aim of unraveling the pathogenesis of MA. Lovastatin, blocks the mevalonate pathway inhibiting the 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-CR), an enzyme of the mevalonate pathway upstream the mevalonate kinase enzyme, reproducing biochemical features similar to those found in MKD.

Journal of Cardiovascular Translational Research, 2013

Since contradictory findings have been reported on potential effects of statins in modulating the... more Since contradictory findings have been reported on potential effects of statins in modulating the inflammatory response, we have analysed the biological activity of lovastatin both in vitro using the Raw 264.7 murine macrophagic cell line and in vivo using BALB/c mice. When added to Raw 264.7 cells in combination with lipopolysaccharide, lovastatin significantly potentiated the release of interleukin-1β, interleukin-6 and interleukin-12 with respect to lipopolysaccharide alone and showed an additive effect on the release of nitric oxide. Similarly, when lovastatin was intraperitoneally administrated to BALB/c mice, it did not induce any proinflammatory effect when used alone, but it significantly potentiated the pro-inflammatory activity of lipopolysaccharide, in terms of number of intraperitoneal cells and serum levels of serum amyloid A, interleukin-1β, interleukin-6 and interleukin-12. A potential clinical implication of our study is that lovastatin might exert a pro-inflammatory activity in subjects affected by inflammatory processes, with clinically evident or subclinical infections.

BioMed Research International, 2013

Mevalonate Kinase Deficiency (MKD) is a rare autosomal recessive inborn disorder of cholesterol b... more Mevalonate Kinase Deficiency (MKD) is a rare autosomal recessive inborn disorder of cholesterol biosynthesis caused by mutations in the mevalonate kinase (MK) gene, leading to MK enzyme decreased activity. The consequent shortage of mevalonate-derived isoprenoid compounds results in an inflammatory phenotype, caused by the activation of the NALP3 inflammasome that determines an increased caspase-1 activation and IL-1 release. In MKD, febrile temperature can further decrease the residual MK activity, leading to mevalonate pathway modulation and to possible disease worsening. We previously demonstrated that the administration of exogenous isoprenoids such as geraniol or the modulation of the enzymatic pathway with drugs, such as Tipifarnib, partially rescues the inflammatory phenotype associated with the defective mevalonic pathway. However, it has not been investigated yet how temperature can affect the success of these treatments. Thus, we investigated the effect of temperature on primary human monocytes from MKD patients. Furthermore the ability of geraniol and Tipifarnib to reduce the abnormal inflammatory response, already described at physiological temperature in MKD, was studied in a febrile condition. We evidenced the role of temperature in the modulation of the inflammatory events and suggested strongly considering this variable in future researches aimed at finding a treatment for MKD.

Pediatric Research, 2008

Mevalonate kinase deficiency (MKD) is a rare disorder characterized by recurrent inflammatory epi... more Mevalonate kinase deficiency (MKD) is a rare disorder characterized by recurrent inflammatory episodes and, in most severe cases, by psychomotor delay. Defective synthesis of isoprenoids has been associated with the inflammatory phenotype in these patients, but the molecular mechanisms involved are still poorly understood, and, so far, no specific therapy is available for this disorder. Drugs like aminobisphosphonates, which inhibit the mevalonate pathway causing a relative defect in isoprenoids synthesis, have been also associated to an inflammatory phenotype. Recent data asserted that cell inflammation could be reversed by the addition of some isoprenoids, such as geranylgeraniol and farnesyl pyrophosphate. In this study, a mouse model for typical MKD inflammatory episode was obtained treating BALB/c mice with aminobisphosphonate alendronate and bacterial muramyldipeptide. The effect of exogenous isoprenoids-geraniol, farnesol, and geranylgeraniol-was therefore evaluated in this model. All these compounds were effective in preventing the inflammation induced by alendronate-muramyldipeptide, suggesting a possible role for these compounds in the treatment of MKD in humans.

Pediatric Research, 2013

Mevalonate kinase deficiency (MKD) is a rare genetic autoinflammatory disease caused by blocking ... more Mevalonate kinase deficiency (MKD) is a rare genetic autoinflammatory disease caused by blocking of the enzyme mevalonate kinase in the pathway of cholesterol and isoprenoids. The pathogenic mechanism originating an immune response in MKD patients has not been clearly understood. We investigated the dysregulation of expression of selected cytokines and chemokines in the serum of MKD patients. The results have been compared with those observed in an MKD mouse model obtained by treating the mice with aminobisphosphonate, a molecule that is able to inhibit the cholesterol pathway, mimicking the genetic block characteristic of the disease. Interleukin (IL)-1β, IL-5, IL-6, IL-9, IL-17, granulocyte colony-stimulating factor, monocyte chemotactic protein-1, tumor necrosis factor-α, and IL-4 expression were dysregulated in sera from MKD patients and mice. Moreover, geraniol, an exogenous isoprenoid, when administered to MKD mice, restored cytokines and chemokines levels with values similar to those of untreated mice. Our findings, which were obtained in patients and a mouse model mimicking the human disease, suggest that these cytokines and chemokines could be MKD specific and that isoprenoids could be considered as potential therapeutic molecules. The mouse model, even if with some limitations, was robust and suitable for routine testing of potential MKD drugs.

Medical Hypotheses, 2012

Crohn&amp... more Crohn's disease (CD) is a multifactorial disease, in which environmental, microbial and genetic factors play important roles. CD is characterized by a chronic granulomatous inflammation by necrotic scarring with aspects of full-thickness wall. In spite of affecting mainly young adults, sometimes, CD can be present in the first year of life (early onset Crohn disease, EOCD) showing an unpredictable course and being often more severe than at older ages. In this paper we propose the hypothesis that EOCD patients should be analyzed using a Mendelian approach with family studies aimed to identify new loci directly involved in the early onset Crohn's disease. So we will leave the classic association study approach used until now for the identification of genes responsible for susceptibility to CD and propose linkage family analysis as alternative and powerful tool for the identification of new genetic variants associated with familiar cases of EOCD.

Medical Hypotheses, 2013

Mevalonate Kinase Deficiency (MKD) is an autosomal-recessively inherited disorder of cholesterol ... more Mevalonate Kinase Deficiency (MKD) is an autosomal-recessively inherited disorder of cholesterol biosynthesis with higher prevalence in the Netherlands and other North European countries. MKD is due to mutations in the second enzyme of mevalonate pathway (mevalonate kinase, MK/MVK) which results in reduced enzymatic activity and in the consequent shortage of downstream compounds. In most severe cases the deregulation of mevalonate pathway is associated with a decrease in serum cholesterol. More than 100 pathological mutations have been described in the MVK gene so far, and a founder effect has been hypothesized as responsible for the diffusion of the most frequent disease-associated mutations. In the acute phase of disease, patients affected with MKD present low cholesterol levels comparable to their basal physiologic conditions, already characterized by lower cholesterol levels when compared to healthy individuals. Low cholesterol levels are widely known to correlate with the reduction of cardiovascular events. We hypothesize a selective advantage for heterozygote carriers of the most frequent MVK mutations in those countries where the diet is characterized by high consumption of saturated animal fats rich in cholesterol. This could explain the maintenance in North European population of the main mutations leading to MKD and the distribution world-wide of these mutations that followed the migrations of North European populations.

International Immunopharmacology, 2010

The inhibition of mevalonate pathway through genetic defects (mevalonate kinase deficiency, MKD) ... more The inhibition of mevalonate pathway through genetic defects (mevalonate kinase deficiency, MKD) or pharmacologic drugs (aminobisphosphonates) causes a shortage of intermediate compounds and, in particular, of geranylgeranyl-pyrophosphate (GGPP) associated to the activation of caspase-1 and IL-1β release. Geraniol (GOH), farnesol (FOH), geranylgeraniol (GGOH) and menthol (MOH), due to their isoprenoid structure, are supposed to enter the mevalonate pathway and to by-pass the biochemical block, reconstituting the pathway. Considering the already known side effects of aminobisphosphonates, and the lack of a specific treatment for MKD, we evaluated the impact of these natural isoprenoids compounds in a RAW cell lines chemically treated with the aminobisphosphonate alendronate, and in monocytes isolated from 2 patients affected by MKD. GOH, FOH, GGOH and MOH were all capable to diminish inflammatory marker levels induced by LPS. These natural isoprenoids could be proposed as novel therapeutic approach for the still orphan drug MKD, but also considered for the evaluation of possible inflammatory side effects of aminobisphosphonates.

Inflammation Research, 2010

Objective The aim of this study was to evaluate, in a mouse model of mevalonate kinase deficiency... more Objective The aim of this study was to evaluate, in a mouse model of mevalonate kinase deficiency (MKD), the possible link between inflammatory symptoms and serum cholesterol levels. Materials and methods Balb/c mice were treated with alendronate and bacterial muramyl dipeptide. Body temperature, interleukin-1b (IL-1b) secretion and serum cholesterol levels were measured. Results An increased production of the pro-inflammatory cytokine IL-1b (p \ 0.05) and a rise in body temperature (p \ 0.05) was observed, while, in parallel, serum cholesterol concentration significantly decreased (p \ 0.05). These effects were completely reversed when animals were treated with exogenous isoprenoids. Conclusions In the mouse model of MKD, the inflammatory response is associated with a reduction in cholesterol levels, and hence this parameter could be used as an indicator of isoprenoid consumption. In addition, plant derived isoprenoids could represent candidate treatments for this disease.

Inflammation Research, 2012

A potential anti-inflammatory effect of clodronate--an aminobisphosphonate--was described to anta... more A potential anti-inflammatory effect of clodronate--an aminobisphosphonate--was described to antagonize the pro-inflammatory effects of the block in the mevalonate pathway, the main feature of a rare auto-inflammatory disease called mevalonate kinase deficiency (MKD). In this study we evaluated the potential anti-inflammatory effect of clodronate in MKD--a still orphan drug pediatric disease. We studied some biological parameters, nitric oxide production using Griess reagents and programmed cell death by flow cytometry, as common inflammatory parameters in MKD, in the presence of different doses of clodronate (1, 10 and 100 μM). In our cellular model and in monocytes from patients with MKD, clodronate induced an increase in programed cell death and nitric oxide production in comparison with non-treated cells. Our findings suggest that clodronate does not have an anti-inflammatory effect as previously reported but that it increases the epiphenomena of this pediatric disease.

Genomics, 2013

The knowledge of the human genome is in continuous progression: a large number of databases have ... more The knowledge of the human genome is in continuous progression: a large number of databases have been developed to make meaningful connections among worldwide scientific discoveries. This paper reviews bioinformatics resources and database tools specialized in disseminating information regarding genetic disorders. The databases described are useful for managing sample sequences, gene expression and post-transcriptional regulation. In relation to data sets available from genome-wide association studies, we describe databases that could be the starting point for developing studies in the field of complex diseases, particularly those in which the causal genes are difficult to identify.

Gene, 2012

Crohn disease (CD) is a multifactorial disorder affecting mainly young adults. Sometimes, however... more Crohn disease (CD) is a multifactorial disorder affecting mainly young adults. Sometimes, however, it can present in the first year of life (Early onset Crohn disease (EOCD)) showing an unpredictable course and can often be more severe than at older ages. Some cases have been associated to an underlying primary immunodeficiency such as IL10R deficiency. We studied the functional response to IL-10 and the genotype of IL-10 receptor in four patients with early onset crohn-like colitis. We found an IL10R variant, which may be associated with a decreased response to the cytokine in one patient. Further studies to determine its pathogenic effect should be performed. In addition IL-10 mediated inhibition of LPS-induced TNFα expression was measured in patient's monocytes.

Apoptosis, 2011

The inhibition of mevalonate pathway by the aminobisphosphonate alendronate (ALD) has been previo... more The inhibition of mevalonate pathway by the aminobisphosphonate alendronate (ALD) has been previously associated with an augmented lipopolysaccharideinduced interleukin-1beta (IL-1b) secretion in monocytes, as demonstrated in an auto-inflammatory disease known as mevalonate kinase deficiency (MKD). In this study we investigated the effect of ALD ? LPS on monocyte cell line (Raw 264.7) death. ALD strongly augmented LPSinduced programmed cell death (PCD) as well as IL-1b secretion in Raw murine monocytes, whereas necrosis was rather unaffected. ALD ? LPS induced caspase-3 activation. Inhibition of IL-1b stimulation partially restored cell viability. These findings suggest that the inhibition of mevalonate pathway, together with a bacterial stimulus, induce a PCD partly sustained by the caspase-3-related apoptosis and partly by caspase-1-associated pyroptosis. The involvement of pyroptosis is a novel hit in our cell model and opens discussions about its role in inflammatory cells with chemical or genetic inhibition of mevalonate pathway.