RESUMEN
BACKGROUND: Surgical repair is recommended for the treatment of high-grade partial and full thickness rotator cuff tears, although evidence shows surgery is not necessarily superior to non-surgical therapy. The purpose of this study was to compare percutaneous orthobiologic treatment to a home exercise therapy program for supraspinatus tears. METHODS: In this randomized-controlled, crossover design, participants with a torn supraspinatus tendon received either 'BMC treatment', consisting of a combination of autologous bone marrow concentrate (BMC) and platelet products, or underwent a home exercise therapy program. After three months, patients randomized to exercise therapy could crossover to receive BMC treatment if not satisfied with shoulder progression. Patient-reported outcomes of Numeric Pain Scale (NPS), Disabilities of the Arm, Shoulder, and Hand, (DASH), and a modified Single Assessment Numeric Evaluation (SANE) were collected at 1, 3, 6, 12, and 24 months. Pre- and post-treatment MRI were assessed using the Snyder Classification system. RESULTS: Fifty-one patients were enrolled and randomized to the BMC treatment group (n = 34) or the exercise therapy group (n = 17). Significantly greater improvement in median ΔDASH, ΔNPS, and SANE scores were reported by the BMC treatment group compared to the exercise therapy group (-11.7 vs -3.8, P = 0.01; -2.0 vs 0.5, P = 0.004; and 50.0 vs 0.0, P < 0.001; respectively) after three months. Patient-reported outcomes continued to progress through the study's two-year follow-up period without a serious adverse event. Of patients with both pre- and post-treatment MRIs, a majority (73%) showed evidence of healing post-BMC treatment. CONCLUSIONS: Patients reported significantly greater changes in function, pain, and overall improvement following BMC treatment compared to exercise therapy for high grade partial and full thickness supraspinatus tears. TRIAL REGISTRATION: This protocol was registered with www. CLINICALTRIALS: gov (NCT01788683; 11/02/2013).
Asunto(s)
Trasplante de Médula Ósea , Estudios Cruzados , Terapia por Ejercicio , Lesiones del Manguito de los Rotadores , Humanos , Masculino , Femenino , Lesiones del Manguito de los Rotadores/terapia , Lesiones del Manguito de los Rotadores/diagnóstico por imagen , Persona de Mediana Edad , Terapia por Ejercicio/métodos , Trasplante de Médula Ósea/métodos , Anciano , Estudios de Seguimiento , Resultado del Tratamiento , Manguito de los Rotadores/diagnóstico por imagen , Manguito de los Rotadores/cirugía , Dimensión del Dolor , Adulto , Medición de Resultados Informados por el Paciente , Imagen por Resonancia MagnéticaRESUMEN
PURPOSE: Knee osteoarthritis (OA) is a common, progressively debilitating joint disease, and the intra-articular injection of autologous bone marrow concentrate (BMC) may offer a minimally invasive method of harnessing the body's own connective tissue progenitor cells to counteract accompanying degenerative effects of the disease. However, the extent to which the progenitor cell content of BMC influences treatment outcomes is unclear. We sought to determine whether patient-reported outcome measures associated with BMC treatment for knee OA are related to the concentration of progenitor cells provided. METHODS: In the present study, 65 patients (72 knees) underwent treatment for knee OA with autologous BMC and self-reported their outcomes for up to one year using follow-up questionnaires tracking function, pain, and percent improvement. A small fraction of each patient's BMC sample was reserved for quantification with a haematological analyzer and cryopreserved for subsequent analysis of potential connective tissue progenitor cells using a colony-forming unit fibroblast (CFU-F) assay. RESULTS: Patients reported significant increases in function and overall percent improvement in addition to decreases in pain relative to baseline levels following treatment with autologous BMC that persisted through 12 months. Patients reporting improved outcomes (46 of 72 knees) received BMC injections having higher CFU-F concentrations than non-responding patients (21.1×103 ± 12.4×103 vs 14.3×103 ± 7.0 x103 CFU-F per mL). A progenitor cell concentration of 18×103 CFU-F per mL of BMC was found to best differentiate responders from non-responders. CONCLUSION: This study provides supportive evidence for using autologous BMC in the minimally invasive treatment of knee OA and suggests that increased progenitor cell content leads to improved treatment outcomes. TRIAL REGISTRATION: ClinicalTrials.gov Identifier: NCT03011398, 1/7/17.
Asunto(s)
Osteoartritis de la Rodilla , Médula Ósea , Trasplante de Médula Ósea/métodos , Humanos , Inyecciones Intraarticulares , Osteoartritis de la Rodilla/cirugía , Dolor/etiología , Medición de Resultados Informados por el Paciente , Células Madre , Resultado del TratamientoRESUMEN
BACKGROUND: Umbilical cord (UC) connective tissues contain plastic-adherent, colony forming unit-fibroblasts (CFU-Fs) amenable to culture expansion for potential therapeutic use. Recently, UC-derived allograft products have been made available to practitioners in orthopaedics and other specialties, by companies purporting "stem cell"-based healing. However, such marketing claims conflict with existing regulations for these human tissues, generating questions over the cellular and protein composition of current commercially available UC allograft products. PURPOSE: To evaluate commercial UC allograft products for viable cells, CFU-Fs, and protein makeup. STUDY DESIGN: Descriptive laboratory study. METHODS: Five commercial UC allograft products claiming to contain viable, undescribed "stem cells," 2 obtained from UC blood (UCB) and 3 from UC tissue (UCT), were analyzed. Image-based methods were used to measure cell concentration and viability, a traditional CFU-F assay was used to evaluate in vitro behavior indicative of a connective tissue progenitor cell phenotype often referred to as mesenchymal stem/stromal cells, and quantitative immunoassay arrays were used to measure a combination of cytokines and growth factors. Bone marrow concentrate (BMC) and plasma derived from the blood and bone marrow of middle-aged individuals served as comparative controls for cell culture and protein analyses, respectively. RESULTS: Viable cells were identified within all 5 UC allograft products, with those derived from UCB having greater percentages of living cells (40%-59%) than those from UCT (1%-22%). Compared with autologous BMC (>95% viability and >300 million living cells), no CFU-Fs were observed within any UC allograft product (<15 million living cells). Moreover, a substantial number of proteins, particularly those within UCB allograft products, were undetectable or present at lower concentrations compared with blood and bone marrow plasma controls. Interestingly, several important growth factors and cytokines, including basic fibroblast growth factor, hepatocyte growth factor, interleukin-1 receptor antagonist, and osteoprotegerin, were most prevalent in 1 or more UCT allograft products as compared with blood and bone marrow plasma. CONCLUSION: CFU-Fs, often referred to as stem cells, were not found within any of the commercial UC allograft products analyzed, and clinicians should remain wary of marketing claims stating otherwise. CLINICAL RELEVANCE: Any therapeutic benefit of current UC allograft products in orthopaedic medicine is more likely to be attributed to their protein composition (UCT > UCB) or inclusion of cells without colony forming potential (UCB > UCT).
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Sangre Fetal , Cordón Umbilical , Aloinjertos , Técnicas de Cultivo de Célula , Células Cultivadas , Ensayo de Unidades Formadoras de Colonias , Humanos , Persona de Mediana EdadRESUMEN
PURPOSE: The prevalence of connective tissue progenitor cells within a cell-based therapy is often quantified using the colony-forming unit fibroblast (CFU-F) assay. The present study investigates the feasibility of using cryopreserved bone marrow aspirate concentrate (BMAC) as an alternative cell source to fresh BMAC for CFU-F quantification. METHODS: Freshly prepared and corresponding cryopreserved BMAC samples from patients receiving autologous cell therapy (n = 98) were analyzed using the CFU-F assay for comparison. Cultures were established by directly plating BMAC at low cell densities and maintained for a 2-week growth period. Colonies were enumerated to determine CFU-F frequency, and a subset of cultures was imaged and analyzed to quantify colony area and density. RESULTS: A nonlinear relationship was observed between plating density and CFU-F frequency over a wide range in plating densities (~30-fold). Cryopreserved BMAC yielded recoverable (77 ± 23%) and viable (73 ± 9%) nucleated cells upon thawing. After cryopreservation, CFU-F frequencies were found to be significantly lower (56.6 ± 34.8 vs. 50.3 ± 31.7 colonies per million nucleated cells). Yet the number of CFU-F in fresh and cryopreserved BMAC were strongly correlated (r = 0.87) and had similar area and densities. Further, moderate correlations were observed between the number of CFU-F and nucleated cells, and both the mean colony area and density were negatively correlated with patient age. Notably, no relationship was found between CFU-F frequency and age, regardless of whether fresh or cryopreserved BMAC was used. CONCLUSIONS: Freshly prepared and cryopreserved BMAC yielded correlated results when analyzed using the CFU-F assay. Our findings support the cryogenic storage of patient BMAC samples for retrospective CFU-F analyses, offering a potential alternative for characterizing BMAC and furthering our understanding of progenitor cells in relation to clinical outcome.
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Células de la Médula Ósea/citología , Médula Ósea/metabolismo , Ensayo de Unidades Formadoras de Colonias/métodos , Criopreservación , Fibroblastos/citología , Adolescente , Adulto , Anciano , Recuento de Células , Núcleo Celular/metabolismo , Supervivencia Celular , Femenino , Humanos , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Células Madre/citología , Adulto JovenRESUMEN
Primary human hepatocytes (PHHs) are a limited resource for drug screening, their quality for in vitro use can vary considerably across different lots, and a lack of available donor diversity restricts our understanding of how human genetics affect drug-induced liver injury (DILI). Induced pluripotent stem cell-derived human hepatocyte-like cells (iPSC-HHs) could provide a complementary tool to PHHs for high-throughput drug screening, and ultimately enable personalized medicine. Here, we hypothesized that previously developed iPSC-HH-based micropatterned co-cultures (iMPCCs) with murine embryonic fibroblasts could be amenable to long-term drug toxicity assessment. iMPCCs, created in industry-standard 96-well plates, were treated for 6 days with a set of 47 drugs, and multiple functional endpoints (albumin, urea, ATP) were evaluated in dosed cultures against vehicle-only controls to enable binary toxicity decisions. We found that iMPCCs correctly classified 24 of 37 hepatotoxic drugs (65% sensitivity), while all 10 non-toxic drugs tested were classified as such in iMPCCs (100% specificity). On the other hand, conventional confluent cultures of iPSC-HHs failed to detect several liver toxins that were picked up in iMPCCs. Results for DILI detection in iMPCCs were remarkably similar to published data in PHH-MPCCs (65% versus 70% sensitivity) that were dosed with the same drugs. Furthermore, iMPCCs detected the relative hepatotoxicity of structural drug analogs and recapitulated known mechanisms of acetaminophen toxicity in vitro. In conclusion, iMPCCs could provide a robust tool to screen for DILI potential of large compound libraries in early stages of drug development using an abundant supply of commercially available iPSC-HHs.
Asunto(s)
Enfermedad Hepática Inducida por Sustancias y Drogas/etiología , Fibroblastos/efectos de los fármacos , Hepatocitos/efectos de los fármacos , Ensayos Analíticos de Alto Rendimiento , Células Madre Pluripotentes Inducidas/efectos de los fármacos , Pruebas de Toxicidad/métodos , Xenobióticos/toxicidad , Células 3T3 , Albúminas/metabolismo , Animales , Biomarcadores/metabolismo , Enfermedad Hepática Inducida por Sustancias y Drogas/metabolismo , Enfermedad Hepática Inducida por Sustancias y Drogas/patología , Técnicas de Cocultivo , Relación Dosis-Respuesta a Droga , Femenino , Fibroblastos/metabolismo , Fibroblastos/patología , Hepatocitos/metabolismo , Hepatocitos/patología , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Células Madre Pluripotentes Inducidas/patología , Masculino , Ratones , Reproducibilidad de los Resultados , Medición de Riesgo , Urea/metabolismo , Xenobióticos/clasificaciónRESUMEN
Drug-induced liver injury (DILI) is a leading cause of drug attrition. Significant and well-documented differences between animals and humans in liver pathways now necessitate the use of human-relevant in vitro liver models for testing new chemical entities during preclinical drug development. Consequently, several human liver models with various levels of in vivo-like complexity have been developed for assessment of drug metabolism, toxicity, and efficacy on liver diseases. Recent trends leverage engineering tools, such as those adapted from the semiconductor industry, to enable precise control over the microenvironment of liver cells and to allow for miniaturization into formats amenable for higher throughput drug screening. Integration of liver models into organs-on-a-chip devices, permitting crosstalk between tissue types, is actively being pursued to obtain a systems-level understanding of drug effects. Here, we review the major trends, challenges, and opportunities associated with development and implementation of engineered liver models created from primary cells, cell lines, and stem cell-derived hepatocyte-like cells. We also present key applications where such models are currently making an impact and highlight areas for improvement. In the future, engineered liver models will prove useful for selecting drugs that are efficacious, safer, and, in some cases, personalized for specific patient populations.
Asunto(s)
Enfermedad Hepática Inducida por Sustancias y Drogas/diagnóstico , Evaluación Preclínica de Medicamentos/métodos , Hepatocitos/fisiología , Hígado/citología , Células Madre/fisiología , Animales , Enfermedad Hepática Inducida por Sustancias y Drogas/prevención & control , Ensayos Analíticos de Alto Rendimiento , Humanos , Dispositivos Laboratorio en un Chip , Miniaturización , Biología de Sistemas , Ingeniería de Tejidos/métodosRESUMEN
UNLABELLED: Induced pluripotent stem cell-derived human hepatocyte-like cells (iHeps) could provide a powerful tool for studying the mechanisms underlying human liver development and disease, testing the efficacy and safety of pharmaceuticals across different patients (i.e., personalized medicine), and enabling cell-based therapies in the clinic. However, current in vitro protocols that rely upon growth factors and extracellular matrices (ECMs) alone yield iHeps with low levels of liver functions relative to adult primary human hepatocytes (PHHs). Moreover, these low hepatic functions in iHeps are difficult to maintain for prolonged times (weeks to months) in culture. Here, we engineered a micropatterned coculture (iMPCC) platform in a multiwell format that, in contrast to conventional confluent cultures, significantly enhanced the functional maturation and longevity of iHeps in culture for at least 4 weeks in vitro when benchmarked against multiple donors of PHHs. In particular, iHeps were micropatterned onto collagen-coated domains of empirically optimized dimensions, surrounded by 3T3-J2 murine embryonic fibroblasts, and then sandwiched with a thin layer of ECM gel (Matrigel). We assessed iHep maturity by global gene expression profiles, hepatic polarity, secretion of albumin and urea, basal cytochrome P450 (CYP450) activities, phase II conjugation, drug-mediated CYP450 induction, and drug-induced hepatotoxicity. CONCLUSION: Controlling both homotypic interactions between iHeps and heterotypic interactions with stromal fibroblasts significantly matures iHep functions and maintains them for several weeks in culture. In the future, iMPCCs could prove useful for drug screening, studying molecular mechanisms underlying iHep differentiation, modeling liver diseases, and integration into human-on-a-chip systems being designed to assess multiorgan responses to compounds.