RESUMO
Activating mutations in the NOTCH1 gene are found in over 50 % of T-ALL cases. Since Notch signaling contributes to the leukemia cell survival and growth, targeting Notch signaling using γ-secretase inhibitors (GSI) has been proposed as a molecularly targeted therapy for the treatment of T-ALL. However, not all T-ALL with NOTCH1 activating mutations respond to GSI treatment. We examined whether GSI could enhance the cytotoxic effect of anti-leukemic agents in the GSI-resistant T-ALL cells although GSI does not have anti-tumor effect as a single agent. GSI significantly increased cell death induced by Vincristine (VCR) but not other anti-leukemic drugs (Methotrexate, Asparaginase, and Cytarabine). The GSI effect in enhancing VCR efficacy was not the result of inhibition of Notch signaling. GSI augmented VCR-induced mitotic arrest, followed by apoptosis. GSI accelerated VCR-triggered loss of mitochondrial membrane potential and caspase-mediated apoptosis. Our finding suggests that GSI has other functions besides inhibiting Notch signaling in T-ALL and incorporating GSI into the conventional regimen containing VCR may offer therapeutic advantage by potentiating VCR treatment in leukemia patients.
Assuntos
Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Leucemia-Linfoma Linfoblástico de Células T Precursoras/patologia , Receptores Notch/metabolismo , Vincristina/farmacologia , Secretases da Proteína Precursora do Amiloide/metabolismo , Caspases/metabolismo , Linhagem Celular Tumoral , Dipeptídeos/farmacologia , Resistência a Medicamentos , Sinergismo Farmacológico , Humanos , Potencial da Membrana MitocondrialRESUMO
The germinal center (GC) is the dynamic microenvironment where Ag-activated B cells rapidly expand and differentiate, generating plasma cells (PC) that produce high affinity antibodies. B cells within the GC have great heterogeneity, containing B cells at different stages of activation and differentiation. However, there are few surface markers that allow subsets of GC-B cells to be distinguished. In the present study, we show that GC-B cells in human tonsils contain two distinct populations regarding CD9 expression; CD9- and CD9+ cells. CD9+ GC-B cells are functionally more differentiated towards PC based upon the following evidence; (1) CD9+ cells express higher levels of PC transcription factor, Blimp-1 while lower levels of B cell transcription factors, Bcl-6 and Pax-5, compared to CD9- cells, (2) CD9+ cells differentiate into plasmablasts faster than CD9- cells in the presence of cytokines that generate PC, and (3) CD9 expression was induced in CD9- GC-B cells under PC generating condition and gradually increased in the course of PC differentiation. Taken together, our data suggest that CD9 is a novel marker for a human GC-B cell subset that is committed to PC lineage.
Assuntos
Subpopulações de Linfócitos B/imunologia , Diferenciação Celular/imunologia , Centro Germinativo/imunologia , Plasmócitos/imunologia , Tetraspanina 29/metabolismo , Biomarcadores/metabolismo , Linhagem da Célula , Células Cultivadas , Centro Germinativo/citologia , Humanos , Tonsila Palatina/citologia , Tonsila Palatina/imunologiaRESUMO
PURPOSE: Cisplatin (CDDP)-based chemotherapy is a first-line treatment for patients with advanced head and neck squamous cell carcinomas (HNSCC), despite a high rate of treatment failures, acquired resistance, and subsequent aggressive behavior. The purpose of this study was to study the mechanism of CDDP resistance and metastasis in HNSCC. We investigated the role of NRF2 pathway activation as a driven event for tumor progression and metastasis of HNSCC. EXPERIMENTAL DESIGN: Human HNSCC cell lines that are highly resistant to CDDP were generated. Clonogenic survival assays and a mouse model of oral cancer were used to examine the impact of NRF2 activation in vitro and in vivo on CDDP sensitivity and development of metastasis. Western blotting, immunostaining, whole-exome sequencing, single-cell transcriptomic and epigenomic profiling platforms were performed to dissect clonal evolution and molecular mechanisms. RESULTS: Implantation of CDDP-resistant HNSCC cells into the tongues of nude mice resulted in a very high rate of distant metastases. The CDDP-resistant cells had significantly higher expression of NRF2 pathway genes in the presence of newly acquired KEAP1 mutations, or via epigenomic activation of target genes. Knockdown of NRF2 or restoration of the wild-type KEAP1 genes resensitized resistant cells to CDDP and decreased distant metastasis (DM). Finally, treatment with inhibitor of glutaminase-1, a NRF2 target gene, alleviated CDDP resistance. CONCLUSIONS: CDDP resistance and development of DM are associated with dysregulated and epigenetically reprogrammed KEAP1-NRF2 signaling pathway. A strategy targeting KEAP1/NRF2 pathway or glutamine metabolism deserves further clinical investigation in patients with CDDP-resistant head and neck tumors.
Assuntos
Antineoplásicos , Neoplasias de Cabeça e Pescoço , Fator 2 Relacionado a NF-E2 , Carcinoma de Células Escamosas de Cabeça e Pescoço , Animais , Humanos , Camundongos , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral , Cisplatino/farmacologia , Cisplatino/uso terapêutico , Resistencia a Medicamentos Antineoplásicos/genética , Epigênese Genética , Epigenômica , Neoplasias de Cabeça e Pescoço/tratamento farmacológico , Neoplasias de Cabeça e Pescoço/genética , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Camundongos Nus , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Transdução de Sinais , Carcinoma de Células Escamosas de Cabeça e Pescoço/tratamento farmacológico , Carcinoma de Células Escamosas de Cabeça e Pescoço/genéticaRESUMO
Adult mesenchymal stromal cells are plastic-adherent cells that are self-renewing and have the capacity to differentiate into various tissue specific lineages. Stromal cells were initially discovered over 100 years ago and substantial insight into stromal cell identification, isolation, characterization, and differentiation has been made, including efforts to elucidate the factors involved in stromal cell differentiation. Stromal cells have immune privilege and thus are attractive candidates for tissue engineering and regenerative medicine applications. Positive results from a number of recent investigations support the use of adult mesenchymal stromal cells for clinical application. This review article provides a brief overview of past, present, and future stromal cell technology.
Assuntos
Células-Tronco Adultas , Células-Tronco Mesenquimais , Medicina Regenerativa/tendências , Adulto , Células-Tronco Adultas/citologia , Células-Tronco Adultas/fisiologia , Animais , Diferenciação Celular , Humanos , Transplante de Células-Tronco Mesenquimais , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Procedimentos Ortopédicos , Pesquisa com Células-Tronco , Células Estromais/citologia , Células Estromais/fisiologia , Terminologia como Assunto , Engenharia TecidualRESUMO
Tumor-associated macrophages (TAMs) have a significant presence in the tumor stroma across multiple human malignancies and are believed to be beneficial to tumor growth. Targeting CSF1R has been proposed as a potential therapy to reduce TAMs, especially the protumor, immune-suppressive M2 TAMs. Additionally, the high expression of CSF1R on tumor cells has been associated with poor survival in certain cancers, suggesting tumor dependency and therefore a potential therapeutic target. The CSF1-CSF1R signaling pathway modulates the production, differentiation, and function of TAMs; however, the discovery of selective CSF1R inhibitors devoid of type III kinase activity has proven to be challenging. We discovered a potent, highly selective, and orally bioavailable CSF1R inhibitor, IACS-9439 (1). Treatment with 1 led to a dose-dependent reduction in macrophages, promoted macrophage polarization toward the M1 phenotype, and led to tumor growth inhibition in MC38 and PANC02 syngeneic tumor models.
Assuntos
Antineoplásicos/uso terapêutico , Benzotiazóis/uso terapêutico , Neoplasias/tratamento farmacológico , Pirimidinas/uso terapêutico , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/antagonistas & inibidores , Antineoplásicos/síntese química , Antineoplásicos/farmacocinética , Benzotiazóis/síntese química , Benzotiazóis/farmacocinética , Estabilidade de Medicamentos , Humanos , Microssomos Hepáticos/metabolismo , Estrutura Molecular , Pirimidinas/síntese química , Pirimidinas/farmacocinética , Relação Estrutura-Atividade , Células THP-1 , Macrófagos Associados a Tumor/efeitos dos fármacosRESUMO
Inhibition of glutaminase-1 (GLS-1) hampers the proliferation of tumor cells reliant on glutamine. Known glutaminase inhibitors have potential limitations, and in vivo exposures are potentially limited due to poor physicochemical properties. We initiated a GLS-1 inhibitor discovery program focused on optimizing physicochemical and pharmacokinetic properties, and have developed a new selective inhibitor, compound 27 (IPN60090), which is currently in phase 1 clinical trials. Compound 27 attains high oral exposures in preclinical species, with strong in vivo target engagement, and should robustly inhibit glutaminase in humans.
Assuntos
Inibidores Enzimáticos/química , Glutaminase/antagonistas & inibidores , Triazóis/farmacocinética , Administração Oral , Animais , Linhagem Celular Tumoral , Cães , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacocinética , Glutaminase/genética , Glutaminase/metabolismo , Meia-Vida , Hepatócitos/citologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Concentração Inibidora 50 , Masculino , Camundongos , Microssomos/metabolismo , Ligação Proteica , Ratos , Ratos Sprague-Dawley , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Relação Estrutura-Atividade , Triazóis/química , Triazóis/metabolismoRESUMO
Src homology 2 domain-containing phosphatase (SHP2) is a phosphatase that mediates signaling downstream of multiple receptor tyrosine kinases (RTK) and is required for full activation of the MAPK pathway. SHP2 inhibition has demonstrated tumor growth inhibition in RTK-activated cancers in preclinical studies. The long-term effectiveness of tyrosine kinase inhibitors such as the EGFR inhibitor (EGFRi), osimertinib, in non-small cell lung cancer (NSCLC) is limited by acquired resistance. Multiple clinically identified mechanisms underlie resistance to osimertinib, including mutations in EGFR that preclude drug binding as well as EGFR-independent activation of the MAPK pathway through alternate RTK (RTK-bypass). It has also been noted that frequently a tumor from a single patient harbors more than one resistance mechanism, and the plasticity between multiple resistance mechanisms could restrict the effectiveness of therapies targeting a single node of the oncogenic signaling network. Here, we report the discovery of IACS-13909, a specific and potent allosteric inhibitor of SHP2, that suppresses signaling through the MAPK pathway. IACS-13909 potently impeded proliferation of tumors harboring a broad spectrum of activated RTKs as the oncogenic driver. In EGFR-mutant osimertinib-resistant NSCLC models with EGFR-dependent and EGFR-independent resistance mechanisms, IACS-13909, administered as a single agent or in combination with osimertinib, potently suppressed tumor cell proliferation in vitro and caused tumor regression in vivo. Together, our findings provide preclinical evidence for using a SHP2 inhibitor as a therapeutic strategy in acquired EGFRi-resistant NSCLC. SIGNIFICANCE: These findings highlight the discovery of IACS-13909 as a potent, selective inhibitor of SHP2 with drug-like properties, and targeting SHP2 may serve as a therapeutic strategy to overcome tumor resistance to osimertinib.
Assuntos
Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Neoplasias Experimentais/patologia , Proteína Tirosina Fosfatase não Receptora Tipo 11/antagonistas & inibidores , Acrilamidas/farmacologia , Compostos de Anilina/farmacologia , Animais , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Receptores ErbB/genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Camundongos , Mutação , Neoplasias Experimentais/genética , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
OBJECTIVE: To compare initial femoral fixation properties of a new implant, Graftgrab (GG), with 2 established methods of extra-articular femoral graft fixation, spiked washers (SW) and bone staples (ST). STUDY DESIGN: Experimental in vitro cohort study. METHODS: Canine semitendinosus-gracilis tendon grafts were passed through bone tunnels and fixed to the lateral surface of femoral condyles with spiked washers, bone staples, or new implant prototypes. The fixations were tested to failure with a single-cycle load at a rate of 50 mm/min. Failure and yield strength, stiffness, energy, and elongation were determined from load-displacement curves and failure modes were recorded. RESULTS: The graft failed midsubstance in 4 SW, 4 ST, and 1 GG fixations. In 3 SW, 3 ST, and 1 GG specimens, the graft slipped from the fixation. The graft ruptured at the clip (3) and the intra-articular (2) surface of the bone tunnel in the remaining GG specimens. There were no significant differences between fixation groups in femoral tunnel length, femoral width, or the mechanical properties evaluated. CONCLUSIONS: The initial in vitro mechanical properties of the new fixation implant are comparable with those of spiked washers and bone staples. CLINICAL RELEVANCE: The initial mechanical performance of the new implant tested in this study was similar to those of comparable, established implants. The new implant is novel and may be useful for human anterior and veterinary cranial cruciate ligament graft reconstruction fixation.
Assuntos
Lesões do Ligamento Cruzado Anterior , Ligamento Cruzado Anterior/cirurgia , Cães/cirurgia , Procedimentos de Cirurgia Plástica/veterinária , Transferência Tendinosa/veterinária , Tendões/transplante , Animais , Fenômenos Biomecânicos , Estudos de Coortes , Procedimentos de Cirurgia Plástica/métodos , Valores de Referência , Transferência Tendinosa/métodosRESUMO
The objective of this study was to determine the tissue density, in vitro expansion and differentiation of canine adipose tissue-derived (ASC) and bone marrow-derived (BMSC) stromal cells. Primary (P0) and cell passages 1-6 (P1-6) cell doubling numbers (CD) and doubling times (DT) were determined in fresh cells. The P0, P3, and P6 adipogenic (CFU-Ad), osteogenic (CFU-Ob), and fibroblastic (CFU-F) colony forming unit frequencies, lineage specific mRNA levels in differentiated P3 cells and composition of P3 and P6 chondrogenic pellets were assessed in cryogenically preserved cells. Cell yields from bone marrow were significantly higher than adipose tissue. Overall ASC and BMSC CDs and DTs and P3 and P6 CFU-F, CFU-Ad, and CFU-Ob were comparable. The P0 BMSC CFU-Ob was significantly higher than ASC. Lineage specific mRNA levels were higher in differentiated versus control cells, but similar between cell types. Protein was significantly greater in P3 versus P6 ASC chondrogenic pellets. Based on these findings, fresh and revitalized canine ASCs are viable alternatives to BMSCs for stromal cell applications.
Assuntos
Tecido Adiposo/citologia , Células da Medula Óssea/citologia , Células Estromais/citologia , Animais , Técnicas de Cultura de Células/veterinária , Diferenciação Celular , Cães , Feminino , MasculinoRESUMO
Mesenchymal stromal cell research has advanced significantly over the last decade, especially in the area of adult adipose-derived stem cells (ASCs). There are significant species differences in ASC harvest, as well as in vitro doubling time and differentiation. Selection of appropriate animal models for preclinical evaluations is critical for optimization and validation of ASC therapeutic effects. As such, in vitro studies are necessary precursors to in vivo ASC applications. Additionally, to elucidate environmental, disease, or trauma effects on native stromal cell populations, characterization of cells from unexposed individuals is necessary. Rats are an established immunocompetent small animal model for a large number of human disorders and toxin exposures in the majority of organ systems. Isolation and expansion techniques for rat ASCs continue to change with advances in technology and ASC biology. The information presented in this chapter is a summary of rat ASC harvest, isolation, quantification, and in vitro expansion.
Assuntos
Tecido Adiposo/citologia , Envelhecimento/fisiologia , Diferenciação Celular , Separação Celular/métodos , Adipogenia , Animais , Proliferação de Células , Células Cultivadas , Condrogênese , Criopreservação , Osteogênese , Ratos , Células Estromais/citologiaRESUMO
Stromal cells are undifferentiated cells found in embryonic and adult tissues. Adult mesenchymal stromal cells (MSCs) possess the properties of self renewal, long-term viability, multipotentiality, and immune privilege, which make them attractive candidates for regenerative medicine applications. In order to develop targeted adult stromal cell therapies for diseased and injured tissues in animals and humans, it is essential to have large-animal models. The dog represents not only a patient population, but is also a valuable experimental model. The dog has contributed significantly to the understanding of various human diseases such as genetic and musculoskeletal disorders. In order to optimize the use of stromal cell therapy in the dog as a patient or disease model, a comprehensive characterization of the cells is required.
Assuntos
Tecido Adiposo/citologia , Envelhecimento/fisiologia , Técnicas de Cultura de Células/métodos , Adipogenia , Animais , Bioensaio , Proliferação de Células , Separação Celular , Células Cultivadas , Condrogênese , Criopreservação , DNA Complementar/biossíntese , Cães , Glicosaminoglicanos/metabolismo , Compostos Orgânicos/metabolismo , Osteogênese , Papaína/metabolismo , RNA/isolamento & purificação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Estromais/citologiaRESUMO
Equine degenerative suspensory ligament desmitis (DSLD) is a debilitating condition that has limited response to rest and stall confinement. This study was designed to test the hypothesis that mild to moderate DSLD is not worsened by consistent exercise. Paso Fino and Peruvian Paso horses (two normal horses and four horses with DSLD) were exercised for 30 min every other day for 8 weeks and then pasture rested for 4 months. Gait analysis, radiographs, ultrasound and serum insulin and glucose concentrations were performed prior to the exercise trial and at each time point. Vertical impulse increased after 8 weeks of exercise and 4 months of pasture rest in DSLD-affected horses. Suspensory ligament fiber pattern subjectively improved with exercise in affected horses. Insulin levels significantly decreased from baseline in all horses after 4 and 8 weeks of exercise. Exercise did not seem to exacerbate and may have improved signs of DSLD in mild to moderate cases.
Assuntos
Terapia por Exercício/veterinária , Doenças dos Cavalos/terapia , Ligamentos/fisiopatologia , Doenças Musculoesqueléticas/veterinária , Condicionamento Físico Animal/fisiologia , Animais , Glicemia/análise , Estudos de Casos e Controles , Feminino , Marcha/fisiologia , Doenças dos Cavalos/fisiopatologia , Cavalos , Insulina/sangue , Coxeadura Animal/fisiopatologia , Coxeadura Animal/terapia , Ligamentos/diagnóstico por imagem , Masculino , Doenças Musculoesqueléticas/fisiopatologia , Doenças Musculoesqueléticas/terapia , Radiografia , Índice de Gravidade de Doença , Resultado do Tratamento , UltrassonografiaRESUMO
It is well established that bone maintenance and healing is compromised in alcoholics. Adult bone marrow-derived stromal cells (BMSCs) and adipose tissue-derived stromal cells (ASCs) likely contribute to bone homeostasis and formation. Direct and indirect alcohol exposure inhibits osteoprogenitor cell function through a variety of proposed mechanisms. The goal of this study was to characterize the effects of chronic alcohol ingestion on the native number and in vitro growth characteristics and multipotentiality of adult BMSCs and ASCs in a rat model. Adult male Sprague-Dawley rats received a liquid diet containing 36% ethanol or an isocaloric substitution of dextramaltose (control). After 4, 8, or 12 weeks of the diet, ASCs were harvested from epididymal adipose tissue and BMSCs from femoral and tibial bone marrow. Cell doublings (CDs) per day and doubling times (DTs) were determined for primary cells (P0) and cell passages 1 through 6 (P1-P6). Fibroblastic (CFU-F), adipogenic (CFU-Ad), and osteogenic (CFU-Ob) colony-forming unit (CFU) frequencies were assessed for P0, P3, and P6. The CDs and DTs were lower and higher, respectively, for ASCs and BMSCs harvested from ethanol versus control rats at all time points. The CFU-F, CFU-Ad, and CFU-Ob were significantly higher in ASCs harvested from control versus ethanol rats for P0, P3, and P6 at all times. Both CFU-Ad and CFU-Ob were significantly higher in P0 BMSCs harvested from control versus ethanol rats after 12 weeks of the diet. The CFU-Ob for P3 BMSCs from control rats was significantly higher than those from ethanol rats after 8 and 12 weeks on the diet. All three CFU frequencies in ASCs from ethanol rats tended to decrease with increasing diet duration. The ASC cell and colony morphology was different between control and ethanol cohorts in culture. These results emphasize the significant detrimental effects of chronic alcohol ingestion on the in vitro expansion and multipotentiality of adult mesenchymal stromal cells (MSCs). Maintenance of the effects through multiple cell passages in vitro suggests cells may be permanently compromised.
Assuntos
Alcoolismo/fisiopatologia , Células da Medula Óssea/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Estromais/efeitos dos fármacos , Tecido Adiposo/citologia , Animais , Células da Medula Óssea/citologia , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Masculino , Osteogênese/efeitos dos fármacos , Osteogênese/fisiologia , Ratos , Ratos Sprague-Dawley , Células-Tronco , Células Estromais/citologiaRESUMO
Colorectal cancer (CRC) is the third most common malignancy and the second leading cause of cancer-related deaths in America. Nearly two thirds of newly diagnosed CRC cases include lymph node (LN) involvement, and LN metastasis is one of the strongest negative prognostic factors for CRC. It is thought that CRC tumors contain a small population of drug-resistant CRC tumor-initiating cells (Co-TICs) that may be responsible for cancer recurrence. To evaluate the effects of the LN stromal cells on Co-TICs, we established a unique xenoplant model using CRC cells isolated by enzymatic digestion from consented patient specimens, HT-29 cells, HCA-7 cells, and LN stromal cell line HK cells. We found that HK cells and HK cell-conditioned media enhanced CRC tumor formation and tumor angiogenesis. Cells expressing CD133(+) and the stromal cell-derived factor 1α (SDF-1α) receptor CXCR4 were enriched in chemotherapeutic-resistant CRC cells. CD133(+)CXCR4(+) Co-TICs isolated from patient specimens are more tumorigenic than unsorted tumor cells. Furthermore, the inhibitors specific to HK cell-derived SDF-1α reduced tumor formation and tumor angiogenesis. Our results have demonstrated a role for Co-TICs in tumor growth and defined the influence of LN stromal cells on Co-TICs. We have identified a major Co-TIC/LN microenvironment-specific mechanism for CRC resistance to chemotherapeutic agents and established experimental platforms for both in vitro and in vivo testing, indicating that SDF-1α and its receptor, CXCR4, may be targets for clinical therapy.
Assuntos
Antígenos CD/biossíntese , Quimiocina CXCL12/metabolismo , Neoplasias do Colo/metabolismo , Glicoproteínas/biossíntese , Linfonodos/citologia , Comunicação Parácrina , Receptores CXCR4/metabolismo , Células Estromais/fisiologia , Antígeno AC133 , Animais , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Neoplasias do Colo/irrigação sanguínea , Neoplasias do Colo/tratamento farmacológico , Resistencia a Medicamentos Antineoplásicos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Transplante de Neoplasias , Neovascularização Patológica , Peptídeos , Transdução de Sinais , Transplante HeterólogoRESUMO
Adipose-derived stem cells (ASCs) express a nonimmunogenic profile as shown by in vitro studies that demonstrate a lack of T cell proliferation to allogeneic ASCs as well as ASC-mediated suppression of mixed lymphocyte reactions. To determine whether these observations would translate in vivo, immune monitoring studies were carried out in conjunction with a rat spinal fusion study. ASCs derived from Fischer or ACI strain rats were loaded onto scaffolds and implanted in Fischer recipients that had undergone the following treatments: (1) No treatment; (2) Scaffold only; (3) Syngeneic ASCs+Scaffold; or (4) Allogeneic ASCs+Scaffold. Half of each group was sacrificed at 4 weeks postimplantation, and the remaining animals were sacrificed at 8 weeks. As determined in a separate study, allogeneic and syngeneic ASCs were equally efficacious in accelerating spinal fusion compared to No treatment and Scaffold only control groups. To determine whether donor ASCs induced an immune response in recipient rats, lymph nodes were harvested for T cell proliferation studies and serum was collected to assess antibody responses. Although T cell priming was not detected to donor alloantigens in recipients at either time point, significant antibody responses were detected to ACI ASCs in animals implanted with syngeneic or allogeneic ASCs. Antibodies were of the IgG isotype, noncytotoxic in the presence of complement, and reactive to fetal bovine serum. These results support the use of allogeneic ASCs for spinal fusion.
Assuntos
Tecido Adiposo/citologia , Fusão Vertebral , Células-Tronco/imunologia , Animais , Anticorpos/metabolismo , Formação de Anticorpos/efeitos dos fármacos , Formação de Anticorpos/imunologia , Proliferação de Células/efeitos dos fármacos , Concanavalina A/farmacologia , Citometria de Fluxo , Tolerância Imunológica , Imunidade Celular/efeitos dos fármacos , Imunidade Celular/imunologia , Modelos Animais , Fenótipo , Ratos , Ratos Endogâmicos F344 , Baço/citologia , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Linfócitos T/citologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Transplante HomólogoRESUMO
Posterolateral spinal fusion is the standard treatment for lumbar compression fractures. Adult adipose tissue-derived stem cells (ASCs) promote osteogenesis in vivo and in vitro. The hypothesis tested in this study was that syngeneic and allogeneic ASCs on a biomaterial scaffold composed of tricalcium phosphate and collagen I will accelerate spinal fusion in a rat model. ASCs from male Fischer or ACI rats were loaded onto scaffolds (53,571 cells/mm(3)) and cultured in stromal media for 48 h. Male Fisher rats were assigned to 4 cohorts (n = 14/cohort) after bilateral decortication of the L4 and L5 transverse processes: (1) No treatment; (2) scaffold only; (3) scaffold + syngeneic ASCs; or (4) scaffold + allogeneic ASCs. Half of each cohort was harvested 4 or 8 weeks after surgery. Spinal fusion was evaluated with radiographs, microcomputed tomography, and light microscopy. Callus did not form in spines without scaffolds. There were no significant differences in callus formation among scaffold cohorts 4 weeks after surgery. Callus formation was more mature in both ASC cohorts versus scaffold alone 8 weeks after surgery based on microstructure as well as radiographic and microcomputed tomographic evidence of active bone formation. Inflammatory cell infiltrate was significantly lower in both ASC cohorts (syngeneic = 18.3 +/- 0.85%; allogeneic = 23.5 +/- 2.33%) versus scaffold alone (46.8 +/- 11.8%) 4 weeks after surgery. Results of this study support syngeneic and allogeneic ASC acceleration of posterior lumbar spinal fusion in a rat model.