Ana Carolina De Faria Morandini

Ana Carolina De Faria Morandini

Education

Education
May 2009 - Present

PhD Candidate

University of São Paulo, Bauru School of Dentistry

Provisional Title:  Study of the role of Toll-like receptors 2 and 4  in cytokines production by gingival and periodontal ligament fibroblasts

Advisor: Dr. Carlos Santos

Abstract: Increasing interest has been directed to the role of the innate immune system in inflammatory diseases, including periodontitis. In essence, several pathogen associated molecular patterns (PAMPs) from bacteria can interact with pattern recognition receptors (PRRs) such as Toll-like receptors (TLRs) and have been found to be important for the innate immune system. The innate host response to lipopolysaccharide (LPS) from Porphyromonas gingivalisis unusual in that different studies have reported that it can be an agonist for TLR2 as well as an antagonist or agonist for TLR4. Besides inflammatory cells, fibroblasts express TLRs and the osteotropic cytokine Interleukin (IL)-6 as well as the chemokine IL-8 are expressed by human gingival fibroblasts (HGF) and human periodontal ligament fibroblasts (HPLF). The present study aim to investigate whether signaling through TLR2 or TLR4 can affect the production of IL-6 and IL-8 in HGF and HPLF from the same volunteer donors. For this, fibroblasts will be isolated and cultured from gingival and periodontal ligament of systemically healthy volunteers and subsequently they will be transfected with a scrambled siRNA, TLR2 siRNA or TLR4 siRNA. After transfection, cells will be challenged with P. gingivalis LPS (1µg/mL), E. coli LPS (1µg/mL), Pam2CSK4 (TLR2/6 agonist, 50 ng/mL), or Pam3CSK4 (TLR2/1 agonist, 500 ng/mL). After 6h, the supernatant will be collected and IL-6 and IL-8 will be analyzed by enzyme-linked immunosorbent assay (ELISA). Quantitative real-time PCR analysis will be performed to confirm the TLR2 and TLR4 Knockdown.

Mar 2007 - Mar 2009

Master of Science

University of São Paulo, Bauru School of Dentistry

Title: Differential production of macrophage inflammatory protein-1alpha, stromal-derived factor-1, and IL-6 by human cultured periodontal ligament and gingival fibroblasts challenged with lipopolysaccharide from P. gingivalis

Advisor: Dr. Carlos Santos

Abstract: Fibroblasts are considered important cells in periodontitis. When challenged by different agents, they respond through the release of cytokines that participate in the inflammatory process. The aim of this study is to evaluate and compare the expression and production of macrophage inflammatory protein (MIP)-1alpha, stromal-derived factor (SDF)-1, and interleukin (IL)-6 by human cultured periodontal ligament and gingival fibroblasts challenged with lipopolysaccharide (LPS) from Porphyromonas gingivalis. METHODS: Fibroblasts were cultured from biopsies of gingival tissue and periodontal ligament of the same donors and used on the fourth passage. After confluence in 24-well plates, the culture medium alone (control) or with 0.1 to 10 microg/ml of LPS from P. gingivalis was added to the wells, and after 1, 6, and 24 hours, the supernatant and the cells were collected and analyzed by enzyme-linked immunosorbent assay and real-time polymerase chain reaction, respectively. RESULTS: MIP-1alpha, SDF-1, and IL-6 protein production was significantly greater in gingival fibroblasts compared to periodontal ligament fibroblasts. IL-6 was upregulated in a time-dependent manner, mainly in gingival fibroblasts, which secreted more MIP-1 alpha in the lowest concentration of LPS used (0.1 microg/ml). In contrast, a basal production of SDF-1 that was inhibited with the  increase of LPS concentration was detected, especially after 24 hours. CONCLUSION: The distinct ability of the gingival and periodontal ligament fibroblasts to secrete MIP-1alpha, SDF-1, and IL-6 emphasizes that these cells may differently contribute to the balance of cytokines in the LPS-challenged periodontium.

2003 - 2006

DDS

University of São Paulo, Bauru School of Dentistry

Title: Concepts and Biological Principles of Periodontal Tissue Engineering - Literature Review

Advisor : Dr. Mario Taba Junior

 *Ana Carolina de Faria Morandini was granted a scholarship (2004-2006) from the Brazilian government, Ministry of Education (MEC/SESu), and was enrolled in the Tutorial Education Program (PET) to perform activities related to Teaching, Research and Community Services. The tutor in this program was Dr. Carlos F. Santos, DDS, PhD, professor from the Discipline of Pharmacology at Bauru School of Dentistry. This scholarship is granted for the top 4 sophomore dental students. In her last year as an undergraduate student, she was selected for participation in an Externship Program in Periodontology at the University of Michigan, School of Dentistry, Ann Arbor, MI in the period of July/August of 2006.

Summary

        Ana Carolina de Faria Morandini graduated from University of São Paulo, Bauru School of Dentistry in 2006. She earned her MS in Oral Rehabilitation/ Periodontics in 2009, followed by the Certificate in Periodontology from Brazilian Federal Board of Dentistry. Currently, she is a PhD candidate in Oral Biology at University of São Paulo, Bauru School of Dentistry and she was granted a scholarship from State of São Paulo Research Foundation (FAPESP 2009/07376-0).

Publications

Morandini AC, Sipert CR, Ramos-Junior ES, Brozoski DT, Santos CF. Periodontal ligament and gingival fibroblasts participate in the production of TGF-β, interleukin (IL)-8 and IL-10. Braz Oral Res. 2011 Apr;25(2):157-62.PMID: 21537641  [PubMed - indexed for MEDLINE]

Abstract: The aim of this study was to quantify and compare the production of transforming growth factor beta (TGF-β), interleukin (IL)-8 and IL-10 by human cultured periodontal ligament and gingival fibroblasts both obtained from the same donors challenged with lipopolysaccharide (LPS) from Porphyromonas gingivalis. Fibroblasts were exposed to 0.1-10µg/mL of LPS from P. gingivalis and after 24 h the supernatants were collected and analyzed by enzyme-linked immunosorbent assay (ELISA). TGF-β protein production was upregulated in a concentration-dependent manner, mainly in gingival fibroblasts, which was statistically significant when challenged by 10µg/mL LPS. Additionally, at this concentration, gingival fibroblasts had almost a two-fold increase in the amount of TGF-βwhen compared to periodontal ligament fibroblasts. Both periodontal ligament and gingival fibroblasts showed an increase in IL-8 production when challenged with 1µg/mL and 10µg/mL LPS. IL-10 production remained unaffected when challenged by any of the LPS concentrations tested in either periodontal ligament or gingival fibroblasts. Our results demonstrate that periodontal ligament and gingival fibroblasts when challenged by LPS from P. gingivalis with 24 h may play a critical role in producing TGF-β and IL-8 but not IL-10.

Morandini AC, Sipert CR, Gasparoto TH, Greghi SL, Passanezi E, Rezende ML, Sant'ana AP, Campanelli AP, Garlet GP, Santos CF. Differential production of macrophage inflammatory protein-1alpha, stromal-derived factor-1, and IL-6 by human cultured periodontal ligament and gingival fibroblasts challenged with lipopolysaccharide from P. gingivalis. J Periodontol. 2010 Feb;81(2):310-7. PMID: 20151811  [PubMed - indexed for MEDLINE]

Abstract: Fibroblasts are considered important cells in periodontitis. When challenged by different agents, they respond through the release of cytokines that participate in the inflammatory process. The aim of this study is to evaluate and compare the expression and production of macrophage inflammatory protein (MIP)-1a, stromal-derived factor (SDF)-1, and interleukin (IL)-6 by human cultured periodontal ligament and gingival fibroblasts challenged with lipopolysaccharide (LPS) from Porphyromonas gingivalis. Methods: Fibroblasts were cultured from biopsies of gingival tissue and periodontal ligament of the same donors and used on the fourth passage. After confluence in 24-well plates, the culture medium alone (control) or with 0.1 to 10 mg/ml of LPS from P. gingivalis was added to the wells, and after 1, 6, and 24 hours, the supernatant and the cells were collected and analyzed by enzyme-linked immunosorbent assay and realtime polymerase chain reaction, respectively. Results: MIP-1a, SDF-1, and IL-6 protein production was significantly greater in gingival fibroblasts compared to periodontal ligament fibroblasts. IL-6 was upregulated in a time-dependent manner, mainly in gingival fibroblasts, which secreted more MIP-1a in the lowest concentration of LPS used (0.1 mg/ml). In contrast, a basal production of SDF-1 that was inhibited with the increase of LPS concentration was detected, especially after 24 hours. Conclusion: The distinct ability of the gingival and periodontal ligament fibroblasts to secrete MIP-1a, SDF-1, and IL-6 emphasizes that these cells may differently contribute to the balance of cytokines in the LPS-challenged periodontium.

Morandini AC, Santos CF, Taba Jr M. Concepts and Biological Principles of Periodontal Tissue Engineering - Literature Review. Revista Peridontia da Sociedade Brasileira de Periodontologia (SOBRAPE) 2008, v. 18, p. 14-18.

Abstract: The main goal of periodontal therapy must be the structural and functional rehabilitation of the lost tissues due to the periodontal disease, resulting in repair and formation of new periodontal attachment (new bone, cementum and periodontal ligament). Regeneration depends on the proliferation, migration, differentiation and production of extracellular protein matrix. Tissue engineering is an emerging multidisciplinary field defined as the application of the concepts of engineering, chemistry and biology aiming repair, restoration or regeneration of tissues. The aim of this literature review is to discuss principles and methods of engineering tissues for periodontal regeneration emphasizing on cell therapy, as well as to show the state of the art in this field and to present the future directions in Dentistry.

Dos Santos Andrade D, Serra RR, Svensjö E, de Araújo Lima AP, Junior ES, da Silva de Azevedo Fortes F, de Faria Morandini AC, Morandi V, de Nazaré Correia Soeiro M, Tanowitz HB, Scharfstein J. "Trypanosoma cruzi invades host cells through the activation of endothelin and bradykinin receptors: a converging pathway leading to chagasic vasculopathy". Br J Pharmacol. 2011 Jul 28. [Epub ahead of print] PubMed PMID: 21797847

Abstract: Independent studies in experimental models of Trypanosoma cruzi appointed different roles for endothelin-1 (ET-1) and bradykinin (BK) in the immunopathogenesis of Chagas disease. Here we addressed the hypothesis that pathogenic outcome is influenced by functional interplay between endothelin receptors (ET(A) R and ET(B) R) and bradykinin receptors (B(2) R). Experimental approach: Intravital microscopy was used to determine whether ETR/B(2) R drives the accumulation of rhodamine-labeled leukocytes in the hamster cheek pouch (HCP). Inflammatory edema was measured in the infected BALB/c paw. Parasite invasion was assessed in CHO overexpressing ETRs, mouse cardiomyocytes, endothelium (HUVECs) or smooth muscle cells (HSMCs), in the presence/absence of antagonists of B(2) R (HOE-140), ET(A) R (BQ-123) and ET(B) R (BQ-788); specific IgG antibodies to each GPCRs; cholesterol or calcium-depleting drugs. RNA interference (ET (A) R or ET(B) R genes) in parasite infectivity was investigated in HSMCs. Key results: BQ-123, BQ-788 and HOE-140 reduced leukocyte accumulation in HCP topically exposed to trypomastigotes and blocked edematogenic inflammation in infected mice. Acting synergistically, ET(A) R and ET(B) R antagonists reduced parasite invasion of HSMCs to the same extent as HOE-140. Exogenous ET-1 potentiated T. cruzi uptake by HSMCs via ETRs/B(2) R whereas RNA interference of ET(A) R and ET(B) R genes conversely reduced parasite internalization. ETRs/B(2) R-driven infection in HSMCs was reduced in HSMC pretreated with MβCD, a cholesterol depleting drug, or in thapsigargin or verapamil treated target cells. Conclusions and Implications: Our findings suggest that plasma leakage, a neutrophil-driven inflammatory response evoked by trypomastigotes via the kinin/endothelin pathways, may offer a window of opportunity for enhanced parasitism of cardiovascular cells.

* Submitted for Publication

Morandini AC, Souza PPC, Ramos-Junior ES, Brozoski, DT, Sipert CR, Souza CAS, Santos CF. Toll-like receptors 2 and 4 knockdown modulates IL-6 and IL-8 in human periodontal fibroblasts

Abstract

Aim: Instead of only playing a structural role in the construction and maintenance of connective tissue, fibroblasts are now seen as active components of the immune response because these cells express Toll-like receptors (TLRs), recognize pathogen associated molecular patterns and mediate the production of cytokines and chemokines during inflammation. This study investigated whether signaling through TLR2 or TLR4 can affect the production of IL-6, IL-8, CXCL12 and CCL3 in human gingival fibroblasts (HGF) and human periodontal ligament fibroblasts (HPLF). Materials and methods: After small interfering RNA-mediated silencing of TLR2 and TLR4, HGF and HPLF were stimulated with Porphyromonas gingivalis lipopolysaccharide (LPS), Escherichia coli LPS, Pam2CSK4 and Pam3CSK4.IL-6, IL-8, CXCL12 and CCL3 mRNA and protein were evaluated by qRT-PCR and ELISA, respectively. Results: Knockdown of TLR2 decreased IL-6 and IL-8 quantities in response to P. gingivalis LPS, Pam2CSK4 and Pam3CSK4 in both fibroblasts subpopulations. During Pam2CSK4 and Pam3CSK4 stimulation knockdown of TLR4 also decreased IL-6. Conversely, quantities of CXCL12 and CCL3 chemokines remained unchanged by TLR2 or TLR4 silencing. Conclusions: These results suggest that signaling through the innate immune system by the most numerous resident cells in periodontium controls the production of important pro-inflammatory cytokines which contribute to periodontal pathogenesis.

Publications in Annals of Events

Pinheiro, CR. ; Sipert, CR. ; Gasparoto, TH. ; Oliveira, C.E. ; Silva, MS. ; Morandini, AC ; Garlet, GP. ; Santos, CF. ; Silva, JS. ; Campanelli, AP. "Toll-like Receptor 4(TLR4) is expressed in Pulp Fibroblasts After Stimulation with Candida Albicans". In: American Association of Endodontists 2011 Annual Session, 2011, San Antonio. American Association of Endodontists 2011 Annual Session, 2011.

Schwartz-Filho, HO ; Wennerberg, A. ; Jimbo, R ; Ramos-Junior, ES. ; Morandini, AC. ; Marcantonio, RAC . Cytokines production by primary cultured gingival fibroblasts under different titanium surfaces. Latin America Osseointegration Congress, São Paulo, SP, 2011

Sipert, CR. ; Morandini, AC; Oliveira, SHP ; Campanelli, AP ; Santos, CF. Differential production of CXCL12 by Cultured human dental pulp and gingival fibroblasts challenged by Porphyromonas gingivalis LPS and Escherichia coli LPS. In: 6th International Conference on Innate Immunity, 2009, Crete. 6th International Conference on Innate Immunity. Crete - Greece : Aegean Conferences, 2009. p. 132-132

Salmeron, S. ; Morandini, AC ; Figueira, EA. ; PassaneziI, AC. ; Rezende, MLR. ; GreghiI, SLA. The use of Platelets Rich Plasm (PRP) in combination with sub-epithelial gingival graft  for root recovery in tabagists: a case report . In: XXI Jornada Odontológica de Bauru, 2008, Bauru. Annals XXI JOB, 2008

Prestes, MP; Machado, MAAM ; Santos, CF. ; Sakai, VT. ; Morandini, AC. Effect of 25% metronidazole gel in plaque index, gingival index and probing depth in type I diabetic children.  In: 23rd Annual SBPqO Meeting, 2006, Atibaia, SP. Brazilian Oral Research. São Paulo, SP : SBPqO, 2006. v. 20. p. 231-231.

Poster Presentations in Conferences

Morandini, AC ; Souza, PPC. ; Ramos-Junior, ES. ; Sipert, CR. ; Costa, CAS. ; Santos, CF. Small Interfering RNA-Mediated Silencing of Toll-like Receptors 2 and 4 Modify Interleukin (IL)-6 and IL-8 Production by Human Gingival and Periodontal Ligament Fibroblasts. Basic Science Research Finalist Poster at 97th Meeting of the American Academy of Periodontology. Miami Beach, Florida,  2011.

Morandini, AC ; Sipert, CR. ; Ramos-Junior, ES. ; Greghi, SLA. ; Campanelli, AP. ; Santos, CF.  Cytokines Produced by Periodontal Fibroblasts after Porphyromonas gingivalis Lipopolysaccharide Challenge. 88th General Session & Exhibition of the IADR, Barcelona, Spain, 2010

Morandini, AC. ; Ferreira Jr, SB ; Repeke, CEP. ; Santos, CF. ; Greghi, SLA. ; Martins Jr, W. ; Silva, JS. ; Campanelli, AP. ; Garlet, GP. IL-6-174 SNP and P gingivalis independently modulate IL-6 expression in periodontitis. 86th General Session & Exhibition of the IADR, Toronto, Canadá,  2008.