INTRODUÇÃO: O paratormônio (PTH) e o fator de crescimento de fibroblastos 23 (FGF-23) aumentam precocemente durante o curso da doença renal crônica (DRC) antes do desenvolvimento de hiperfosfatemia. Este estudo avaliou os efeitos de dois quelantes de fósforo, acetato de cálcio (Ca) e hidrocloreto de sevelamer (SEV), nos níveis de PTH e FGF-23 de pacientes com DRC. MÉTODOS: Quarenta e dois pacientes com DRC estágios III e IV foram randomizados em 2 grupos para receber durante 6 semanas, Ca ou Sev. Após este período os pacientes foram seguidos por mais 2 semanas (washout). Analisamos os efeitos destes quelantes sobre os parâmetros do metabolismo ósseo e mineral. RESULTADOS: No início do estudo, os pacientes apresentaram-se com fração de excreção do fósforo, PTH e FGF-23 séricos elevados. Durante o tratamento com quelantes de fósforo houve um declínio progressivo nos níveis de PTH e fósforo urinário, mas sem mudanças nos níveis séricos de cálcio e fósforo. Ocorreu uma mudança significativa nos níveis de FGF-23 no grupo de pacientes tratados com Sev. CONCLUSÕES: Este estudo confirmou os efeitos positivos da prescrição de quelantes de fósforo no controle do PTH, nos estágios III e IV da DRC. Estudos prospectivos e de longo seguimento são necessários para confirmar os efeitos do Sev sobre os níveis de FGF-23 e os benefícios de sua redução sobre parâmetros como mortalidade / INTRODUCTION: Parathyroid hormone (PTH) and fibroblast growth factor (FGF-23) levels increase early in CKD before the occurrence of hyperphosphatemia. This study evaluated the effect of two phosphate binders, calcium carbonate or sevelamer hydrocloride, on PTH and FGF-23 levels in patients with CKD. METHODS: Forty two patients were randomized in two groups to receive calcium acetate or sevelamer hydrochloride, over a 6-wk period. After that, the patients were followed by more two weeks and effects of phosphate binders on mineral parameters were analyzed. RESULTS: At baseline, patients presented with elevated fractional excretion of phosphate, serum PTH and FGF-23 During treatment with both phosphate binders, there was a progressive decline in serum PTH and urinary phosphate, but no change in serum calcium or serum phosphate. Significant changes were observed for FGF-23 only in sevelamer-treated patients. CONCLUSIONS: This study confirms the positive effects of early prescription of phosphate binders on PTH control. Prospective and long-term studies are necessary to confirm the effects of sevelamer hydrocloride on serum FGF-23 and the benefits of this decrease on outcomes

Base teórica: A artrite reumatoide (AR) é uma doença autoimune sistêmica onde a inflamação crônica da sinóvia articular e a subsequente erosão óssea e da cartilagem resultam em destruição articular, dor e incapacidade funcional. Apesar dos recentes progressos no tratamento da AR, estes ainda apresentam limitações e significativos efeitos adversos, salientando a necessidade de novas estratégias terapêuticas. Plantas da família das Amarilidáceas apresentam em seus bulbos um conjunto de alcaloides muito característicos e exclusivos com atividades farmacológicas, tais como atividade antiviral, anti-inflamatória e atividade anticolinérgica. A montanina é um alcaloide isolado da Rhodophiala bífida, uma planta da família das amarilidáceas utilizada na medicina popular, mas nunca antes testada como terapia para doenças inflamatórias. Objetivo: Avaliar o efeito da montanina como uma terapia anti-inflamatória in vivo em dois modelos de artrite e in vitro sobre a proliferação de linfócitos e sobre a invasão de fibroblastos sinoviais (FLS). Métodos: Artrite induzida por antígeno (AIA) foi realizada em camundongos Balb/C com albumina bovina sérica metilada e a nocicepção e a migração de leucócitos para a articulação do joelho foram os parâmetros avaliados. Artrite induzida por colágeno (CIA) foi realizada em camundongos DBA/1J e o desenvolvimento e severidade da artrite foi avaliado através de escore clínico, nocicepção articular e escore histológico. Montanina foi administrada via intraperitoneal, duas vezes ao dia. A proliferação de linfócitos estimulados por concanavalina A (conA) foi realizada pelo método de MTT e invasão de FLS em 24 horas foi avaliada em um sistema de insertos de Matrigel. Resutados: A administração de montanina diminuiu a migração articular de leucócitos (p0,001) e a nocicepção (p0,01) em camundongos com AIA. Em camundongos com CIA, o tratamento com montanina reduziu a severidade da artrite e o dano articular avaliado pelos escores clínico (p0,01) e histológico (p0,05) e melhorou a nocicepção articular (p0,05), sem causar nenhum dano hepático. Além disso, montanina inibiu in vitro a proliferação de linfócitos estimulados com conA (p0,01) e diminuiu a invasão de FLS (p0,05) em 54%, com uma ação independente de citotoxicidade. Conclusão: Esses resultados indicam que a montanina pode ser explorada para se tornar um possível fármaco para o tratamento de doenças inflamatórias e autoimunes, como a AR. / Background: Rheumatoid arthritis (RA) is an autoimmune and systemic disease where the chronic inflammation of articular synovia and the subsequent bone and cartilage erosion results in joint destruction, pain and functional disability. Despite recent progress in RA treatments, its still have limitations and significant side effects, emphasizing the need of new therapeutic strategies. Amaryllidaceae plants presenting at its bulbs a set of very characteristics and exclusives alkaloids with pharmacological activities such as antiviral, anti-inflammatory and anticholinergic activity. Montanine is an alkaloid isolated from the Rhodophiala bifida, an Amaryllidaceae plant used in alternative medicine but never before tested as a therapy for inflammatory diseases. Objective: To evaluate the effect of montanine as an in vivo anti-inflammatory therapy in two arthritis models and in vitro on lymphocytes proliferation and fibroblast-like synoviocytes (FLS) invasion. Methods: Antigen-induced arthritis (AIA) was performed in Balb/C mice with methylated bovine serum albumin and nociception and leukocytes migration into the knee joint were evaluated. Collagen-induced arthritis (CIA) was performed in DBA/1J mice and arthritis development and severity were assessed by clinical scoring, articular nociception and histological scoring. Montanine was administered intraperitoneally twice a day. Lymphocyte proliferation stimulated by concanavalin A in 48 hours was performed with MTT assay and FLS invasion in 24 hours was assayed in a Matrigel-coated transwell system. Results: Administration of montanine decreased leukocyte articular migration (p0.001) and nociception (p0.01) in mice with AIA. In mice with CIA, treatment with montanine reduced severity of arthritis and joint damage assessed by clinical (p0.01) and histological score (p0.05) and ameliorates articular nociception (p0.05), without causing any hepatic damage. Moreover, montanine inhibited in vitro lymphocyte proliferation stimulated with ConA (p0.01) and decreased FLS invasion by 54% (p0.05), with an action independent of cytotoxicity. Conclusion: These findings suggest that montanine can be explored to become a possible medicament to treat inflammatory and autoimmune diseases such as arthritis.

Uma premissa da Biologia Molecular atual estabelece a ativação de programas de transcrição voltados a processos biológicos específicos. Neste trabalho o objetivo é descrever genes regulados que, uma vez agrupados, são capazes de indicar os programas disparados na linhagem corticoadrenal murina Y1 quando tratada com o fator de crescimento de fibroblasto 2 (FGF2) ou pelo hormônio adrenocorticotrópico (ACTH). O papel de ACTH em células Y1 ainda não está bem estabelecido quanto ao seu potencial mitogênico, uma vez que este hormônio é capaz de agir por diferentes vias de sinalização, apresentando um comportamento dual. Ao traçar o perfil de genes regulados por este hormônio, comparando com o padrão observado por tratamentos de indutores clássicos, espera-se determinar o papel de ACTH frente ao ciclo celular. FGF2 é classicamente conhecido por sua atividade mitogênica, de forma que, em células Y1, é capaz de induzir a passagem G0?G1?S do ciclo celular. Recentemente foi descrita uma nova e surpreendente ação de FGF2, como um indutor de morte seletivo, agindo apenas em células potencialmente tumorigênicas (Costa e Armelin, dados não publicados). Foram feitos estudos de expressão gênica com ensaios de microarray, observando-se o padrão de transcrição da linhagem Y1, bem como de sub-linhagens de Y1 resistentes a morte desencadeada por FGF2, submetidas a tratamentos de FGF2, ACTH e soro. Os resultados indicam que a) o padrão de expressão gênica observado quando a linhagem Y1 é submetida a tratamentos de ACTH é diferente daqueles desencadeados por mitógenos clássicos; b) o tratamento de FGF2 regula genes envolvidos na via de MAPK, controle do ciclo celular e genes relacionados a processos de adesão, comunicação intercelular e sinalização a partir da matriz extracelular (ECM); c) o comportamento de morte induzida por FGF2 está relacionado a alterações na estrutura da célula, envolvendo mecanismos de adesão, remodelagem de citoesqueleto e transdução de sinais a partir da ECM. / Nowadays, a molecular biology premise establishes the activation of transcription programs related to specific biological processes. At this work the objective is to describe regulated genes that, once clustered, are able to indicate the running programs when murine adrenocortical lineage Y1 is treated with fibroblast growth factor (FGF2) or by adrenocorticotropin hormone (ACTH). The mitogenic potential of ACTH treatments in Y1 cells is not well established since this hormone acts by different signaling pathways, presenting a dual behavior. By tracing the profile of genes regulated by this hormone and comparing it to the transcription patterns observed in response to classic mitogens it is expected to determine the ACTH role in the cell cycle. FGF2 is known by its mitogenic activity, inducting the G0? G1?S cell cycle transitions in Y1 cells. Recently it has been described a new and surprising feat of FGF2, acting as a selective death inductor, only in potentially tumorigenic cells (Costa and Armelin, unpublished data). Microarray assays were used to determine the transcription patterns observed in Y1 lineage, as well as in FGF2 death-resistant Y1 sub-lineages, submitted to FGF2, ACTH and serum treatments. The results indicate that a) the gene expression profile displayed when Y1 cells are under ACTH treatments is different from the patterns observed when this lineage is submitted to classic mitogens; b) FGF2 treatment regulates genes involved in the MAPK pathway, cell cycle control and genes related to adhesion processes, intercellular communication and signaling from the extra cellularmatrix (ECM); c) the death behavior initiated by FGF2 is related to structural alterations in the cell, involving adhesion mechanisms, citoskeleton remodeling and signal transduction from the ECM.

Pentoxifilina (PTF), um derivado da metilxantina, tem ação terapêutica como agente antifibrótico. In vitro, a PTF causa inibição na produção de colágeno, glicosaminoglicanos e fibronectina, bem como promove acentuada redução na proliferação dos próprios fibroblastos de quelóides. A PTF na concentração de 1.000 g/mL foi seletiva no controle da inibição da síntese protéica pelos fibroblastos. O objetivo deste estudo foi verificar o comportamento in vitro de fibroblastos oriundos de cicatrizes hipertróficas (HSHF) e de pele não-cicatricial (NHF) na presença e ausência de PTF (1.000 g/mL), quanto à: proliferação celular, produção de colágeno e capacidade dos fibroblastos gerarem contração em modelo experimental de matriz de colágeno. Para tanto, foram utilizados fibroblastos cultivados a partir de amostras de cicatrizes hipertróficas e pele não-cicatricial doadas, com finalidade de pesquisa, pelo banco de Tecidos do Instituto Central do Hospital das Clínicas da FMUSP. Culturas celulares expostas a PTF apresentaram diminuição na proliferação celular em HSHF (46,35%) e em NHF (37,73%) (p<0,0001). Na presença de PTF, foi observada seletividade de inibição na síntese de colágenos, havendo inibição mais expressiva de colágeno tipo III em HSHF e de colágeno tipo I em NHF (p<0,0001). O modelo experimental de matriz de colágeno povoada por fibroblastos de cicatriz hipertrófica apresentou contração menor (12%) na presença de PTF (p<0,0001), em relação à sua ausência / Fibroblasts are thought to be partially responsible for the persisting contractile forces that result in burn contractures. Using a monolayer and fibroblast populated collagen lattice (FPCL) three-dimensional (3D) model we subjected hypertrophic scar and non-cicatricial fibroblasts to the antifibrogenic agent pentoxifylline (PTF) 1000g/mL attempting to reduce proliferation, collagen type I and III synthesis and contraction in this 3D model. Fibroblasts were isolated from post burn hypertrophic scars (HSHF) and non-scarred skin (NHF). Cells were grown in monolayer or incorporated into FPCL\'s and exposed to PTF. In monolayer, cell number proliferation was reduced (46.35% in HSHF group and 37.73% in NHF group) p<0,0001. The PTF also demonstrated to be selective on collagen type I and III synthesis inhibition suggesting higher inhibition of collagen type III on HSHF group and more evident inhibition of type I on NHF group. FPCL\'s containing PTF had surface areas reduced in about 12% p<0,0001. PTF showed inhibition effects on cell proliferation and reduced contraction in both HSHF and NHF

Many physiological and pathological processes, such as wound healing and tissue remodeling, are heavily influenced by continuous mechanical cell-cell and cell-ECM communication. Abnormalities that may compromise the biomechanical communication between the cells and the ECM can have significant repercussions on these physiological and pathological processes. The state of the mechanical environment and the reciprocal communication of mechanical signals between the ECM and the cell during wound healing and aged dermal tissue regeneration may be key in controlling the quality of the structure and physical properties of regenerated tissue. This dissertation encompasses a series of studies developed for characterizing the effects of mechanical cues on altering and controlling tissue remodeling, and regeneration in the context of controlling scar formation during wound healing, and the maintenance and regeneration of the dermal extracellular matrix (ECM) during aging. In order to achieve this goal, in vitro models that contained some features of the provisional ECM, and the ECM of the dermis were developed and subjected to an array of quantifiable mechanical cues. Wound models were studied with different mechanical boundary conditions, and found to exhibit differences in initial short-term structural remodeling that lead to significant differences in the long-term synthesis of collagen after four weeks in culture. Dermal models seeded with fibroblasts from individuals of different ages were treated with a hyaluronic acid (HA)-based dermal filler. Changes in the mechanical environment of the dermal models caused by swelling of the hydrophilc HA, resulted in changes in the expression of mechanosensitive, and ECM remodeling genes, essential for the maintenance and regeneration of dermal tissue. Taken together, these data provide new insights on the role of mechanical signals in directing tissue remodeling.

There exists a dire need for improved therapeutics to achieve predictable and effective bone regeneration. Non-viral gene therapy is a safe method that can efficiently transfect target cells, therefore is a promising approach to overcoming the drawbacks of protein delivery of growth factors. The goal of this study was to employ cost-effective biomaterials to deliver genetic materials (DNA or RNA) in a controlled manner in order to address the high cost issues, safety concerns, and lower transfection efficiencies that exist with protein and gene therapeutic approaches. To achieve our goal, we set several aims: 1) To assess the bone regeneration capacity of polyethylenimine (PEI)-chemically modified ribonucleic acid (cmRNA) (encoding bone morphogenetic protein-2 (BMP-2)) activated matrices, compared to PEI-plasmid DNA (BMP-2)-activated matrices. 2) To explore the osteogenic potential of cmRNA-encoding BMP-9, in comparison to cmRNA-encoding BMP-2. 3) To use collagen membranes as integral components of a guided bone regeneration protocol and to enhance the bioactivity of collagen membranes by incorporating plasmid DNA (pDNA) or cmRNA encoding bone morphogenetic protein-9 (BMP-9). 4) To test whether the delivery of pDNA encoding BMP-2 (pBMP-2) and fibroblast growth factor-2 (pFGF-2) together can synergistically promote bone repair in a leporine model of diabetes mellitus, a condition that is known to be detrimental to union. 5) To investigated whether there is a synergistic effect on bone regeneration following delivery of pBMP-2 and pFGF-2, insulin and/or vitamin D. These investigations together provided new insights regarding the appropriate treatment methods for patients with fractures. Here we develop and test a non-viral gene delivery system for bone regeneration in challenging animal models utilizing a scaffold carrying PEI-nucleic acid complexes. We utilized three kinds of pDNA encoding either BMP-2, BMP-9 or FGF-2 as well as two kinds of cmRNA encoding either BMP-2 or BMP-9 formulated into PEI complexes. The fabricated nanoplexes were assessed for their size, charge, in vitro cytotoxicity, and capacity to transfect human bone marrow stromal cells (BMSCs). The in vivo functional potency of different nanoplexes embedded in scaffolds was evaluated using a calvarial bone defect model in rats, diaphyseal long bone radial defects in a diabetic rabbit model and intramuscular implantation in a diabetic rat. The results indicate that our non-viral gene delivery system induced migration and differentiation of resident cells to enhance bone regeneration. Together these findings suggest that scaffolds loaded with non-viral vectors harboring cmRNA or pDNA encoding osteogenic proteins may be a powerful tool for stimulating bone regeneration with significant potential for clinical translation.

Tumorigenesis is often caused by the dysregulation of developmental pathways that are activated during repair, a process that recapitulates development. The Hedgehog (Hh) pathway is a signalling pathway essential for cell patterning and identity during embryogenesis. Activation of Hh signalling has been reported in small cell lung cancer progression, but the role of the Hh receptor, Patched1 (Ptch1), remains poorly understood. Therefore, it is imperative that we understand how Ptch1 is involved in development and tissue repair in order to understand its roles in cancer. This project aimed to study the role of Ptch1 during the branching process of lung development and in the regeneration of airway epithelial cells. A conditional knockout approach was utilised to excise Ptch1 by crossing Ptch1 conditional mice with Dermo1-Cre mice (Dermo1Cre+/-;Ptch1lox/lox), thereby activating the Hh pathway in the mesenchyme, independent of ligand. Dermo1Cre+/-;Ptch1lox/lox embryos died at E12.0 and showed secondary lung branching arrest leading to lobe formation defects. Expression of Ptch1, Gli1 and Foxf1 were shown to be upregulated in both proximal and distal lung mesenchyme, indicating inappropriate pathway activation and disruption of the Hh gradient. Fgf10 expression was spatially reduced in Dermo1Cre+/-;Ptch1lox/lox lungs and the addition of Fgf10 to these lungs in culture showed partial restoration of branching, thus Hh signalling was shown to regulate branching via Fgf10. Due to the patterning defect associated with our in vivo model, we took an in vitro approach to delete Ptch1 in lung explants cultures. This also showed reduced branching and validated that mesenchymal proliferation was enhanced after Ptch1 deletion, consistent with the previously reported role of Hh signalling in mesenchymal cell survival. Small cell lung cancer originates in the proximal lung and has been linked to aberrant repair processes. Therefore, Hh signalling in proximal airway repair was investigated. Ptch1 expressing cells were detected in the bronchial epithelium and stroma during homeostasis. But these cells were not detected following polidocanol-induced injury in the murine nasal septum and lung. However during naphthalene-induced repair, Ptch1 expressing cells were detected in the regenerating bronchial epithelium, suggesting that Hh dependent progenitors respond specifically to naphthalene-induced damage and perhaps are pulmonary neuroendocrine or variant Clara cells. Therefore, this project has provided insight into how Ptch1 patterns lung branching and lobe specification during development and also highlights the importance of Ptch1 in pulmonary epithelial regeneration.

A 700bp portion of the promoter of CYP2D25, the porcine microsomal vitamin D 25-hydroxylase was isolated and sequenced. The computer analysis of the sequence revealed the existence of a putative VDRE at 220 bp upstream of the transcription start site. A CYP2D25 promoter-luciferase reporter plasmid was constructed in order to study the transcriptional regulation of the gene. Treatment with the vitamin D metabolites calcidiol and calcitriol suppressed the promoter, provided that the nuclear receptors VDR and RXR were overexpressed. Phenobarbital was also capable of suppressing the promoter if the nuclear receptors PXR or CAR were overexpressed. The 25-hydroxylases are not expressed solely in liver but in a wide array of other organs as well. It is therefore possible at least in theory to study the vitamin D 25-hydroxylation in human subjects using cells from extrahepatic organs, from which biopsy retrieval is easier than from the liver. Dermal fibroblasts are frequently used to study different pathological conditions in human subjects and they are easy to come by. Dermal fibroblasts were shown to express two vitamin D 25-hydroxylases: CYP27A1 and CYP2R1. The expression pattern of CYP2R1 displayed considerable interindividual variation. The fibroblasts were also capable of measurable vitamin D 25-hydroxylation, which makes dermal fibroblasts a possible tool in studying vitamin D 25-hydroxylation in human subjects. Little is known about the regulation of expression and activity of the human vitamin D 25-hydroxylases. Therefore dermal fibroblasts – expressing CYP2R1 and CYP27A1 – and human prostate cancer LNCaP cells, that express CYP2R1 and CYP2J2, were treated with calcitriol and phenobarbital and efavirenz, two drugs that give rise to vitamin D deficiency. Treatment decreased the mRNA levels of CYP2R1 and CYP2J2 provided that the treated cells also expressed the necessary nuclear receptors. CYP27A1 did not respond to any of the treatments. The treatments also managed to decrease the 25-hydroxylating activity of the cells. The results show that vitamin D 25-hydroxylases can be regulated by both endogenous and xenobiotic compounds.

<p>Specification and differentiation of a cell is accomplished by changing its gene expression profiles. These processes require temporally and spatially regulated transcription factors (TFs), to induce the genes that are necessary to a specific cell type. In each cell a set of TFs interact with each other or activate their targets; as development progresses, transcription factors receive regulatory inputs from other TFs and a complex gene regulatory network (GRN) is generated. Adding complexity, each TF can be regulated not only at the transcriptional level, but also by translational, and post-translational mechanisms. Thus, understanding a developmental process requires understanding the interactions between TFs, signaling molecules and target genes which establish the GRN.</p><p>In this thesis, two genes, FoxN2/3, a TF and FGFR1, a component of the FGF signaling pathway are investigated. FoxN2/3 and FGFR1 have different mechanisms that function in sea urchin development; FoxN2/3 regulates gene expression and FGFR1 changes phosphorylation of target proteins. However, their ultimate goals are the same: changing the state of an earlier GRN into the next GRN state. </p><p>First, we characterize FoxN2/3 in the primary mesenchyme cell (PMC) GRN. Expression of foxN2/3 begins in the descendants of micromeres at the early blastula stage; and then is lost from PMCs at the mesenchyme blastula stage. foxN2/3 expression then shifts to the secondary mesenchyme cells (SMCs) and later to the endoderm. Here we show that, Pmar1, Ets1 and Tbr are necessary for activation of foxN2/3 in the descendants of micromeres. The later endomesoderm expression is independent of the earlier expression of FoxN2/3 in micromeres and independent of signals from PMCs. FoxN2/3 is necessary for several steps in the formation of larval skeleton. A number of proteins are necessary for skeletogenesis, and early expression of at least several of these is dependent on FoxN2/3. Furthermore, knockdown (KD) of FoxN2/3 inhibits normal PMC ingression. PMCs lacking FoxN2/3 protein are unable to join the skeletogenic syncytium and they fail to repress the transfating of SMCs into the skeletogenic lineage. Thus, FoxN2/3 must be present for the PMC GRN to control normal ingression, expression of skeletal matrix genes, prevention of transfating, and control fusion of the PMC syncytium.</p><p>Second, we show that the FGF-FGFR1 signaling is required for the oral-aboral axis formation in the sea urchin embryos. Without FGFR1, nodal is induced in all of the cells at the early blastula stage and this ectopic expression of nodal requires active p38 MAP kinase. The loss of oral restriction of nodal expression results in the abnormal organization of PMCs and the larval skeleton; it also induces ectopic expression of oral-specific genes and represses aboral-specific genes. The abnormal oral-aboral axis formation also affected fgf and vegf expression patterns; normally these factors are expressed in two restricted areas of the ectoderm between the oral and the aboral side, but when FGFR1 is knocked down, Nodal expands, and in response the expression of the FGF and VEGF ligands expands, and this in turn affects the abnormal organization of larval skeleton.</p> / Dissertation

Cardiac myofibroblasts participate in post-myocardial infarct (MI) wound healing, infarct scar formation, and remodeling of the ventricle remote to the site of infarction. The role of intracellular calcium handling in cardiac myofibroblasts as a modulator of cellular motility, contractile responses, and proliferation is largely unexplored. We have investigated the role of sodium calcium exchange (Na Ca exchange or NCX1.1) and non-selective cation channels (NSCCs) in regulation of myofibroblast function using a pharmacological inhibitor approach in vitro. Primary myofibroblasts were stimulated with PDGF-BB and cellular chemotaxis, contraction and proliferative responses were characterized using standard bioassays (Costar Transwell apparatuses, pre-formed collagen type I gel deformation assays, and 3H-thymidine incorporation). Stimulated cellular responses were compared to those in the presence of AG1296 (PDGFβR inhibitor), KB-R7943 (NCX inhibitor), gadolinium, nifedipine or ML-7. Immunofluorescence was used to determine localized expression of αSMA, SMemb, NCX1.1, and Cav1.2a in cultured myofibroblasts. Motility of myofibroblasts in the presence of PDGF-BB was blocked with AG1296 treatment. Immunoblotting and immunocytochemical studies revealed expression of NCX1.1 in fibroblasts and myofibroblasts. Motility (in the presence of either PDGF-BB or CT-1), contraction (in the presence of either PDGF-BB or TGFβ1), and proliferation (in the presence of PDGF-BB) were sensitive to KB-R7943 treatment of cells (7.5 and 10 μM for motility, 5 and 10 μM for contractility, and 10 μM for proliferation). Proliferation (in the presence of PDGF-BB), and contractility (in the presence of either PDGF-BB or TGFβ1) but not motility (in the presence of PDGF-BB) are sensitive to nifedipine treatment, while gadolinium treatment was associated only with decreased motility of cells (in the presence of either PDGF-BB, CT-1, or LoFGF-2). We found that ML-7 treatment inhibited cellular chemotaxis, and contraction. Thus cellular chemotaxis, contractile, and proliferation responses were sensitive to different pharmacologic treatment. Regulation of transplasmalemmal calcium movements may be important in cytokine and growth factor receptor-mediated cardiac myofibroblast motility, contractility, and proliferation. Furthermore, our results support the hypothesis that activation of specific calcium transport proteins is an important determinant of physiologic responses. / May 2006