Comparison among bone marrow mesenchymal stem and mononuclear cells to promote functional recovery after spinal cord injury in rabbits

Abstract Purpose: To investigate the efficacy of allogeneic mesenchymal stem-cells and autologous mononuclear cells to promote sensorimotor recovery and tissue rescue. Methods: Female rabbits were submitted to the epidural balloon inflation method and the intravenous cells administrations were made after 8 hours or seven days after injury induction. Sensorimotor evaluation of the hindlimbs was performed, and the euthanasia was made thirty days after the treatment. Spinal cords were stained with hematoxylin and eosin. Results: Both therapies given 8 hours after the injury promoted the sensorimotor recovery after a week. Only the group treated after a week with mononuclear cells showed no significant recovery at post-injury day 14. In the days 21 and 28, all treatments promoted significant recovery. Histopathological analysis showed no difference among the experimental groups. Our results showed that both bone marrow-derived cell types promoted significant sensorimotor recovery after injury, and the treatment made at least a week after injury is efficient. Conclusion: The possibilities of therapy with bone marrow-derived cells are large, increasing the therapeutic arsenal for the treatment of spinal cord injury.

Lisado plaquetario y células mononucleares autólogas de la periodontitis crónica. Presentación de un caso / Autologous platelet lysate and mononuclear stem cell of chronic periodontitis. Case presentation

Introducción: El tratamiento de la periodontitis crónica se prolonga por persistencia de la inflamación en tejidos afectados. La medicina regenerativa muestra resultados alentadores. Es la primera vez en Cuba, según bibliografía revisada, que se usa la combinación de lisado plaquetario y células mononucleares autólogas en este tratamiento. Objetivo: Mostrar el efecto de la combinación del lisado plaquetario y células mononucleares autólogas en el tratamiento de la periodontitis. Presentación de un caso: Paciente femenina de 44 años de edad, con diagnóstico de periodontitis, quien desde hace 1 año lleva tratamiento, sin efectuarse procedimientos quirúrgicos por la inflamación persistente. Es remitida a la consulta de periodoncia del Hospital Enrique Cabrera para recibir tratamiento con medicina regenerativa. Se consideró tratar con lisado de plaquetas autólogas, una vez concluido el raspado y alisador radicular, se prefundieron las encías afectadas con el lisado plaquetario. A los 7 días de aplicado el lisado se constatan las encías sin signos clínicos de inflamación, y disminución ostensible del número de dientes afectados y la profundidad o eliminación de las bolsas periodontales y movilidad dentaria. Al mes se trataron quirúrgicamente los defectos óseos con implante de células mononucleares. Al evaluar a los 6 meses, se hallaron de forma variable, signos radiográficos de organización del trabeculado, definición de la cortical y formación ósea en zona de defectos óseos. Conclusiones: El tratamiento combinado del lisado plaquetario autólogo y células mononucleares muestra una evaluación satisfactoria en el tratamiento de la periodontitis crónica y reduce el tiempo de tratamiento(AU)

Introduction: Treatment of chronic periodontitis is prolonged by persistent inflammation in affected tissues. Regenerative medicine shows encouraging results. It is the first time in Cuba, according to the reviewed literature, that the combination of platelet lysate and autologous mononuclear cells is used in this treatment. Objective: to show the effect of the combination of platelet lysate and autologous mononuclear cells in the treatment of periodontitis. Presentation of the case: A 44-year-old female patient diagnosed with periodontitis who had been treated for 1 year without surgical procedures due to persistent inflammation. She is referred to the periodontics consultation of the Dr. Enrique Cabrera General Hospital to receive treatment with regenerative medicine. Treatment with autologous platelet lysate was considered, once the scaling and root planer had been completed, the affected gingivae were prefixed with the platelet lysate. At 7 days after the lysate is applied, the gingiva shows no clinical signs of inflammation, and a noticeable reduction in the number of affected teeth and the depth or elimination of periodontal pockets and tooth mobility. Bone defects with a mononuclear cell implant were surgically treated one month later. When evaluating at 6 months, radiographic signs of trabecular organization, definition of the cortical bone, and bone formation in the area of bone defects were found variably. Conclusions: The combined treatment of autologous platelet lysate and mononuclear cells shows a satisfactory evaluation in the treatment of chronic periodontitis and reduces treatment time(AU)

Caracterización fenotípica de células madre mesenquimales humanas de médula ósea y tejido adiposo. Resultados preliminares / Phenotypic characterization of mesenchymal human stem cell from bone marrow and adipose tissue. preliminary results

Introducción:las células madre mesenquimales (CMM) poseen características fenotípicas y funcionales que les confieren un amplio potencial terapéutico por su posible uso en la terapia celular regenerativa, en el rechazo del trasplante alogénico y en enfermedades inflamatorias crónicas. Objetivo: evaluar la expresión de moléculas de membrana que permiten identificar la expresión de patrones moleculares característicos de CMM humanas mantenidas en cultivo. Métodos: se estudió la expresión fenotípica de células mononucleares procedentes de médula ósea obtenidas mediante aspiración medular, separadas por gradiente de Ficoll y cultivadas ex vivo entre los pases o subcultivos 3 y 16; y adipocitos cultivados procedentes de la extracción enzimática de tejido adiposo de donantes sanos. Se realizó doble marcaje para las moléculas CD34/CD45, CD34/CD90, CD34/CD117, y CD34/CD44. Resultados: en los resultados preliminares obtenidos se observó que las células cultivadas procedentes de médula ósea, entre los pases 4 y 8 de cultivo expresaron 45,13 por ciento de células CD34-/CD45- (doblemente negativas), lo que correspondió con el 25,24 por ciento de células CD34-/CD90+ y el 96,90 por ciento de CD34-/CD117-. En las células procedentes de cultivo de adipocitos se observó el 52,3 por ciento de CD34-/CD45- (doblemente negativas),12,31 por ciento de CD34-/CD90+,43,31 por ciento de CD34-/CD117- y 64,68 por ciento de CD34-/CD44+.Estos resultados sugieren que ambos cultivos se diferenciaron a CMM. Las CMM procedentes de adipocitos mostraron el 64,68 por ciento de células con expresión de la molécula de adhesión CD44 a la que se atribuyen propiedades funcionalescomo el asentamiento tisular. Conclusiones: estos resultados preliminares permiten corroborar que ambos métodos experimentales de cultivo son efectivos para la obtención de CMM con fines terapéuticos

Introduction:mesenchymal stem cells (MSCs) have phenotypic and functional characteristics whichgives them a broad therapeutic potential for possible use in regenerative cell therapy, allogeneic transplant rejection and chronic inflammatory diseases. Objective: to evaluate the expression of moleculemembranes expression to identify molecular patterns characteristic of human MSCs maintained in culture. Methods:the phenotypic expression of mononuclear cells from bone marrow wereobtained by bone marrow aspiration, separated by Ficoll and cultured ex vivo between passages or subcultures 3 and 16 and adipocytes cultured obtained from enzyme extraction of adipose tissue of healthy donor. Double staining was performed for molecules CD34/CD45, CD34/CD90, CD34/CD117 and CD34/CD44. Results:preliminary results showed that cultured mononuclear cells from bone marrow between passage 4 and 8 of culture expressed 45,13 percent CD34-/CD45- cells (double-negative), corresponding to 25,24 percent CD34-/CD90+ cells and 96,90 percent of CD34-/CD117-. Adipocytes from culture cells showed 52,3 percent CD34-/CD45- (double-negative), 12,31 percent cells CD34-/CD90+, 43,31 percent CD34-/CD117- (double-negative). Our results suggest that both cultures were differentiated to MSCs. Adipocytes from MSCs showed 64,68 percent of cells with expression of CD44 adhesion molecule conferring functional homing properties Conclusions:these preliminary results corroborate that the experimental methods used in cultivation are effective for obtaining MSCs with therapeutic purposes

Impacto del tratamiento con células madre adultas en la osteoartrosis de la rodilla / Impact of the treatment with adult stem cells in knee osteoarthrosis

Introducción: la osteoartrosis de la rodilla (OAR) es la causa más frecuente de artritis en la población por encima de los 55 años de edad. A menudo se asocia con discapacidad y deterioro variable de la calidad de vida. En la actualidad, el tratamiento con células madre ha abierto una alternativa más dentro del arsenal terapéutico con que se cuenta. Objetivo: evaluar la eficacia y seguridad del autotrasplante de células madre adultas en el tratamiento de la osteoartrosis de rodilla. Métodos: se realizó un estudio desde mayo 2009 hasta diciembre 2011, en el que se incluyeron 123 pacientes adultos con OAR atendidos en la consulta de medicina regenerativa, que no respondían a los tratamientos convencionales. Se les realizó el implante intrarticular de células mononucleares de sangre periférica (CMN-SP) autólogas, movilizadas mediante el factor estimulante de colonias granulocíticas. En todos los casos prevalecía el dolor y la dificultad a la marcha al inicio del tratamiento. Resultados: a los seis meses se observó mejoría clínica y radiológica en 117 pacientes (95,1 por ciento) y solo en seis enfermos (4,9 por ciento) no hubo la respuesta deseada, coincidiendo estos con los que presentaron trastornos de alineación marcados, en los cuales el dolor no desapareció, pero sí existió modificación de la intensidad. Conclusiones: el implante de CMN-SP es un método factible, simple, seguro y menos costoso para el tratamiento de OAR. Se recomienda que si existe deformidad angular acentuada marcada, esta debe corregirse antes del implante para que el proceder sea exitoso

Introduction: osteoarthrosis an every day more frequent degenerative disease affecting the quality of life of people over 55 years old and its treatment is a great challenge. The use of stem cells has open a new alternative in the available therapeutic arsenal. Objective: to assess the efficacy and safety of autologous adult stem cell transplantation in the treatment of knee osteoarthrosis. Methods: the study took place from May 2009 to December 2011 in 123 adult patients with OAR from the regenerative medicine outpatient service who had no response to conventional treatments. Intra-articular implantation of autologous peripheral blood mononuclear cells (MNC-SP) mobilized by granulocyte colony stimulating factor was performed. In all cases pain and gait difficulty at baseline prevailed. Results: after six months, clinical and radiological improvement in 117 patients (95,1 percent) were observed and only in six patients (4,9 percent) the expected response was not obtained. These last patients presented marked alignment disorders and the pain did not disappear, but there was a positive change in its intensity. Conclusions: the CMN-SP implant is a feasible, simple, safe and less expensive method for the treatment of knee osteoarthrosis. It is recommended that in cases with marked angular deformity, this disorder should be corrected prior to MNC-SP implantation so the procedure be successful

Transplantation of mononuclear cells from human umbilical cord blood promotes functional recovery after traumatic spinal cord injury in Wistar rats

Cell transplantation is a promising experimental treatment for spinal cord injury. The aim of the present study was to evaluate the efficacy of mononuclear cells from human umbilical cord blood in promoting functional recovery when transplanted after a contusion spinal cord injury. Female Wistar rats (12 weeks old) were submitted to spinal injury with a MASCIS impactor and divided into 4 groups: control, surgical control, spinal cord injury, and one cell-treated lesion group. Mononuclear cells from umbilical cord blood of human male neonates were transplanted in two experiments: a) 1 h after surgery, into the injury site at a concentration of 5 x 10(6) cells diluted in 10 µL 0.9 percent NaCl (N = 8-10 per group); b) into the cisterna magna, 9 days after lesion at a concentration of 5 x 10(6) cells diluted in 150 µL 0.9 percent NaCl (N = 12-14 per group). The transplanted animals were immunosuppressed with cyclosporin-A (10 mg/kg per day). The BBB scale was used to evaluate motor behavior and the injury site was analyzed with immunofluorescent markers to label human transplanted cells, oligodendrocytes, neurons, and astrocytes. Spinal cord injury rats had 25 percent loss of cord tissue and cell treatment did not affect lesion extension. Transplanted cells survived in the injured area for 6 weeks after the procedure and both transplanted groups showed better motor recovery than the untreated ones (P < 0.05). The transplantation of mononuclear cells from human umbilical cord blood promoted functional recovery with no evidence of cell differentiation.

Visualização in vivo de células mononucleares de medula óssea na microcirculação coronariana após isquemia miocárdica / In vivo visualization of bone marrow mononuclear cells in coronary microcirculation after myocardial ischemia

A terapia celular para doenças cardiovasculares representa uma nova e promissora opção terapêutica, principalmente para a cardiopatia isquêmica. A injeção de células mononucleares de medula óssea (CMMO) pela via intracoronariana (ICO) é a mais estudada em ensaios clínicos. Embora já se tenha documentado efeitos benéficos com essa terapia, dados relativos aos mecanismos de interação entre as células transplantadas e o ambiente microvascular cardíaco são escassos. A avaliação das CMMO na microcirculação miocárdica tem o potencial de esclarecer seus mecanismos de ação, abrindo novas possibilidades terapêuticas. O objetivo do estudo foi descrever e avaliar uma inovadora técnica de injeção ICO direta e visualização in vivo das CMMO na microcirculação miocárdica de pequenos roedores, após um período de isquemia miocárdica global (IMG) seguida de reperfusão, na presença e na ausência da aterosclerose. Materiais e Métodos: A técnica de transplante cardíaco heterotópico cervical (TCHC) em murinos foi modificada com a inserção e fixação de um microcateter conectado a uma seringa, no tronco braquiocefálico do animal doador. IMG foi induzida ocluindo-se a artéria nutridora do enxerto com um microclamp vascular, por 1 hora. As CMMO foram isoladas e marcadas antes da injeção ICO. Microscopia intravital de fluorescência (MIF) foi usada para análise da microcirculação coronariana do ventrículo direito (VD), incluindo arteríolas, capilares e vênulas pós-capilares. Parâmetros de perfusão e permeabilidade microvasculares e as interações entre as CMMO e células endoteliais foram estudados. O impacto da aterosclerose na recuperação, fenótipo e função celular também foi avaliado. Resultado: A MIF permitiu análise detalhada da microcirculação coronariana e da cinética das CMMO injetadas pela via ICO. A IMG afetou a microcirculação, reduzindo a densidade capilar funcional (DCF). Tal redução foi maior na presença de aterosclerose...

Cell therapy for cardiovascular disease represents a promising new therapeutic option, especially for ischemic heart disease. The intracoronary (ICo) injection of bone marow mononuclear cells (BMMC) is the most commonly studied rote in clinical trials. Although it has been documented beneficial effects with this therapy, data pertaining the mechanisms of interaction between transplanted cells and cardiac microvascular environment are lacking. Assessement of BMMC in myocardial microcirculation has the potential to clarify its mechanisms of action, opening new therapeutic possibilities. The objective of the study was to describe and to evaluate an innovative technique of direct ICo injection and in vivo visualization of BMMC in myocardial small rodents, after a period of global cardiac ischemia (GMI) and reperfusion, in the presence and absence of atherosclerosis. Materials and Methods: The syngeneic murine cervical heterotopic heart transplantation (CHHT) technique was modified by inserting and fixing a catheter connected to a syringe into the donor animal brachiocephalic trunk. GMI was induced occluding the nutritive artery of the graft for 1 hour. Bone marrow mononuclear cells were isolated and labeled prior to ICo injection. Intravital fluorescence microscopy (IFM) was used for analysis of the coronary microcirculation of the right ventricle (RV), including arterioles, capillaries and venules. Parameters of microvascular perfusion, microvascular permeability and the interactions between the BMMC and endothelial cells were studied. The impact of atherosclerosis on the recovery, phenotype and cellular function was also evaluated. Results: The IFM has allowed detailed analysis of both the coronary microcirculation, as the kinetics of BMMC. The GMI affected microcirculation, reducing the functional capillary density (FCD). This reduction was higher in the presence of atherosclerosis. The main area of cell retention was the capillary network...

Human Umbilical Cord Blood Mononuclear Cell Transplantation in Rats with Intrinsic Sphincter Deficiency

To evaluate the effectiveness of the human umbilical cord blood (HUCB) transplantation for the treatment of intrinsic sphincter deficiency (ISD), we analyzed the short term effects of HUCB mononuclear cell transplantation in rats with induced-ISD. ISD was induced in rats by electro-cauterization of periurethral soft tissue with HUCB mononuclear cell injection after 1 week. The sphincter function measured by mean leak point pressure was significantly improved in the experimental group compared to the control group at 4 weeks. (91.75+/-18.99 mmHg vs. 65.02+/-22.09 mmHg, P=0.001). Histologically, the sphincter muscle was restored without damage while in the control group it appeared markedly disrupted with atrophic muscle layers and collagen deposit. We identified injected HUCB cells in the tissue sections by Di-I signal and Prussian blue staining. HUCB mononuclear cell injection significantly improved urethral sphincter function, suggesting its potential efficacy in the treatment of ISD.

Transplante de Células-tronco marcadas para o miocárdio de paciente com doença de chagas / Labeled stem cells transplantation to the myocardium of a patient with chagas' disease

O transplante de células-tronco é uma nova terapia com objetivo de produzir regeneração cardíaca pela diferenciação ou aumento dos miócitos cardíacos ou proliferação neovascular em pacientes no estágio final de insuficiência cardíaca congestiva secundária a cardiomiopatia dilatada¹, mas os resultados são desconhecidos2,3.

Stem cell transplantation is a new therapy applied to produce cardiac regeneration through differentiation or increase of heart myocytes or neovascular proliferation in patients in the end stage of congestive heart failure secondary to dilated cardiomyopathy¹, but the results are still unknown2,3.