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1.
Journal of Experimental Hematology ; (6): 170-178, 2023.
Article in Chinese | WPRIM | ID: wpr-971120

ABSTRACT

OBJECTIVE@#To investigate the expression of pyruvate kinase M2 (PKM2) in bone marrow mesenchymal stem cells (BMSCs) in myeloma bone disease (MBD) and its effect on osteogenic and adipogenic differentiation of BMSCs.@*METHODS@#BMSCs were isolated from bone marrow of five patients with multiple myeloma (MM) (MM group) and five with iron deficiency anemia (control group) for culture and identification. The expression of PKM2 protein were compared between the two groups. The differences between osteogenic and adipogenic differentiation of BMSCs were assessed by using alkaline phosphatase (ALP) and oil red O staining, and detecting marker genes of osteogenesis and adipogenesis. The effect of MM cell line (RPMI-8226) and BMSCs co-culture on the expression of PKM2 was explored. Functional analysis was performed to investigate the correlations of PKM2 expression of MM-derived BMSCs with osteogenic and adipogenic differentiation by employing PKM2 activator and inhibitor. The role of orlistat was explored in regulating PKM2 expression, osteogenic and adipogenic differentiation of MM-derived BMSCs.@*RESULTS@#Compared with control, MM-originated BMSCs possessed the ability of increased adipogenic and decreased osteogenic differentiation, and higher level of PKM2 protein. Co-culture of MM cells with BMSCs markedly up-regulated the expression of PKM2 of BMSCs. Up-regulation of PKM2 expression could promote adipogenic differentiation and inhibit osteogenic differentiation of MM-derived BMSCs, while down-regulation of PKM2 showed opposite effect. Orlistat significantly promoted osteogenic differentiation in MM-derived BMSCs via inhibiting the expression of PKM2.@*CONCLUSION@#The overexpression of PKM2 can induce the inhibition of osteogenic differentiation of BMSCs in MBD. Orlistat can promote the osteogenic differentiation of BMSCs via inhibiting the expression of PKM2, indicating a potential novel agent of anti-MBD therapy.


Subject(s)
Humans , Adipogenesis , Bone Diseases/metabolism , Bone Marrow Cells , Cell Differentiation , Cells, Cultured , Mesenchymal Stem Cells/physiology , Multiple Myeloma/metabolism , Orlistat/pharmacology , Osteogenesis/genetics
2.
Protein & Cell ; (12): 202-216, 2023.
Article in English | WPRIM | ID: wpr-982531

ABSTRACT

Although the mTOR-4E-BP1 signaling pathway is implicated in aging and aging-related disorders, the role of 4E-BP1 in regulating human stem cell homeostasis remains largely unknown. Here, we report that the expression of 4E-BP1 decreases along with the senescence of human mesenchymal stem cells (hMSCs). Genetic inactivation of 4E-BP1 in hMSCs compromises mitochondrial respiration, increases mitochondrial reactive oxygen species (ROS) production, and accelerates cellular senescence. Mechanistically, the absence of 4E-BP1 destabilizes proteins in mitochondrial respiration complexes, especially several key subunits of complex III including UQCRC2. Ectopic expression of 4E-BP1 attenuates mitochondrial abnormalities and alleviates cellular senescence in 4E-BP1-deficient hMSCs as well as in physiologically aged hMSCs. These f indings together demonstrate that 4E-BP1 functions as a geroprotector to mitigate human stem cell senescence and maintain mitochondrial homeostasis, particularly for the mitochondrial respiration complex III, thus providing a new potential target to counteract human stem cell senescence.


Subject(s)
Humans , Mesenchymal Stem Cells/physiology , Cellular Senescence , Homeostasis , Cell Cycle Proteins/metabolism , Adaptor Proteins, Signal Transducing/metabolism , Mitochondria/metabolism , Electron Transport Complex III/metabolism , Cells, Cultured
3.
Rev. Círc. Argent. Odontol ; 80(231): 19-23, jul. 2022. ilus
Article in Spanish | LILACS | ID: biblio-1392286

ABSTRACT

En el campo de la odontología, prevalecen actualmente alternativas terapéuticas con una filosofía conservadora. Sin embargo, con el advenimiento de los tratamientos con células madre (CM), se amplían las posibilidades terapéuticas, que buscan la combinación y el equilibrio entre la intervención tradicional y las posibilidades de reposición de estructuras anatómicas dañadas, a través de la regeneración de tejidos utilizando células madre o sus derivados (AU)


In the dentistry field, therapeutic alternatives with a conservative philosophy currently prevail. However, with the advent of stem cell (SC) treatments, therapeutic possibilities are expanding, seeking a combination and balance between traditional intervention and the pos- sibility of replacing damaged anatomical structures through tissue regeneration, using stem cells or their derivatives (AU)


Subject(s)
Humans , Stem Cells , Tissue Engineering , Mesenchymal Stem Cells/physiology , Periodontal Ligament/physiology , Regeneration/physiology , Tooth/cytology , Tooth Germ/physiology , Biocompatible Materials/therapeutic use , Bone Regeneration/physiology , Dental Pulp/physiology , Tissue Scaffolds , COVID-19/therapy
4.
Int. j. morphol ; 38(5): 1463-1472, oct. 2020. graf
Article in English | LILACS | ID: biblio-1134463

ABSTRACT

SUMMARY: The vomeronasal organ (VNO) is an accessory organ involved on the olfactory pathway, that detects pheromones and emits signals in order to modulate social and reproductive behavior. The VNO stem cells replace neurons throughout life. The aim of this study was to isolate and characterize cells derived from the vomeronasal organ from New Zealand rabbits. Five male rabbits with 120 days were used for cell isolation and culture. Results: VNO-derived cells presented labelling for proliferation (PCNA), undifferentiated profile (Nanog), neuronal (GFAP), mesenchymal stem cells (CD73, CD90 and CD105 and Stro-1). Also, presence of cytoskeletal (Vimentin, b-tubulin and CK-18) and absence of hematopoietic markers (CD34, CD117 and CD45) both by immunofluorescence and flow cytometry. By PCR it was possible to verify the expression of some undifferentiated profile (Oct-4), neuronal (Nestin) and mesenchymal (CD73, CD105 and Vimentin) genes. Functionally, VNO-derived cells differentiate in vitro into adipocytes, osteocytes and chondrocytes, and presented no tumorigenic potential when injected to Balb/c nu/nu mice. In conclusion, the rabbit VNO-derived cells have a profile that could be supportive to VNO olfactory/neuroreceptor epithelium by delivering factors to epithelial turnover or even by differentiation into epithelial cells to replacement of commissural epithelium.


RESUMEN: El órgano vomeronasal (OVN) es un órgano accesorio de la vía olfatoria, que detecta feromonas y emite señales que afectan la modulación del comportamiento social y reproductivo. Las células madre OVN reemplazan las neuronas durante toda la vida. El objetivo de este estudio fue aislar y caracterizar células derivadas del órgano vomeronasal de conejos raza Nueva Zelanda. Para el aislamiento y el cultivo celular se utilizaron cinco conejos machos con una edad de 120 días. Las células del OVN presentaron etiquetado para la proliferación (PCNA), un perfil indiferenciado (Nanog), neuronal (GFAP), células madre mesenquimales (CD73, CD90 y CD105 y Stro-1). Además, se ob- servó presencia de citoesqueleto (Vimentina, β-tubulina y CK-18) y ausencia de marcadores hematopoyéticos (CD34, CD117 y CD45) tanto por inmunofluorescencia como por citometría de flujo. Me- diante PCR fue posible verificar la expresión de algunos genes de perfil indiferenciado (Oct-4), neuronal (Nestin) y mesenquimatoso (CD73, CD105 y Vimentin). Las células derivadas del OVN se diferencian in vitro en adipocitos, osteocitos y condrocitos, y no presentan un potencial tumorigénico al ser infiltrados en ratones Balb / c nu / nu. En conclusión, las células derivadas de OVN de conejo tienen un perfil que podría ser compatible con el epitelio olfatorio / neurorreceptor de OVN transmitiendo factores al recambio epitelial o incluso mediante la diferenciación en células epiteliales para reemplazar el epitelio comisural.


Subject(s)
Animals , Rabbits/anatomy & histology , Vomeronasal Organ/cytology , Mesenchymal Stem Cells/physiology , Olfactory Bulb/cytology , Stem Cells/physiology , Olfactory Mucosa/cytology , Polymerase Chain Reaction , Fluorescent Antibody Technique , Flow Cytometry , Neurons/physiology
5.
Rev. cuba. hematol. inmunol. hemoter ; 36(2): e1133, abr.-jun. 2020.
Article in Spanish | CUMED, LILACS | ID: biblio-1149903

ABSTRACT

La terapia celular basada en células mesenquimales/estromales se aplica ampliamente en la medicina moderna, aun cuando no todos los mecanismos de supervivencia y diferenciación están identificados. Sin embargo, hace pocos años se comenzaron a encontrar elementos extracelulares que generan nuevos paradigmas. En el presente trabajo se explican las principales características y funciones atribuidas a los exosomas, nanopartículas constituidas por microvesículas secretadas por las células con efecto en la matriz extracelular, y su repercusión como alternativa hacia una medicina regenerativa libre de células. Estas estructuras participan de forma notoria y crucial en la comunicación intercelular, lo que ha supuesto un cambio en el concepto de las funciones y el papel que desempeñan estas vesículas en los organismos vivos, en particular en la restauración de tejidos dañados y la respuesta inflamatoria e inmunológica. Se comentan algunos ejemplos de la repercusión biotecnológica de los exosomas en empresas y el mercado biofarmaceútico(AU)


Mesenchymal/stromal cell ;based therapy is widely applied in modern medicine, even though not all survival and differentiation mechanisms are identified. However, a few years ago, extracellular elements began to be found that generate new paradigms. The present work explains the main characteristics and functions attributed to exosomes, nanoparticles made up of microvesicles secreted by with an effect on the extracellular matrix, and their impact as an alternative towards cell-free regenerative medicine. These structures participate, notoriously and critically, in intercellular communication, which has led to a change in the concept of the functions and role that these vesicles play within living organisms, particularly in the restoration of damaged tissues and the inflammatory and immunological response. Some examples of the exosomes' biotechnological impact on companies and the biopharmaceutical market are discussed(AU)


Subject(s)
Humans , Male , Female , Regenerative Medicine/methods , Exosomes/physiology , Mesenchymal Stem Cells/physiology
6.
Einstein (Säo Paulo) ; 18: eAO5236, 2020. graf
Article in English | LILACS | ID: biblio-1133772

ABSTRACT

ABSTRACT Objective To follow the expansion of mesenchymal stem cells from umbilical cords by two classic senescence markers, p16 (INK4A) and p21 (CDKN1A), using practical, fast, and less expensive methods than the gold standard Western blotting technique, to evaluate its applicability in the laboratory. Methods Mesenchymal stem cells from umbilical cords were isolated from Wharton's jelly and, after quality control, morphological and immunophenotypic characterization by flow cytometry, were expanded in culture until coming close to cell cycle arrest (replicative senescence). Results A comparison was made between young cells, at passage 5, and pre-senescent cells, at passage 10, evaluating the protein expression of the classic cell senescence markers p16 and p21, comparing the results obtained by Western blotting with those obtained by flow cytometry and indirect immunofluorescence. Conclusion Follow-up of cell cultures, through indirect p16 immunofluorescence, allows the identification of mesenchymal stem cells from umbilical cord cultures at risk of reaching replicative senescence.


RESUMO Objetivo Acompanhar a expansão de células-tronco mesenquimais de cordão umbilical por dois marcadores clássicos de senescência, p16 (INK4A) e p21 (CDKN1A), usando métodos práticos, rápidos e com custo menor do que a técnica padrão-ouro de Western blotting, para avaliar sua aplicabilidade em laboratório. Métodos Células-tronco mesenquimais de cordão umbilical foram isoladas da geleia de Wharton e, após controle de qualidade e caracterização morfológica e imunofenotípica por citometria de fluxo, foram expandidas em cultura, até chegarem próximas à parada do ciclo celular (senescência replicativa). Resultados Foi feita a comparação entre células jovens, na passagem 5, e células pré-senescentes, na passagem 10, avaliando a expressão proteica dos marcadores clássicos de senescência celular p16 e p21, comparando os resultados obtidos por Western blotting com os obtidos por citometria de fluxo e imunofluorescência indireta. Conclusão O seguimento de culturas celulares, por meio da imunofluorescência indireta de p16, permite identificar as culturas de células-tronco mesenquimais de cordão umbilical em risco de atingirem a senescência replicativa.


Subject(s)
Humans , Umbilical Cord/physiology , Fluorescent Antibody Technique/methods , Cellular Senescence , Mesenchymal Stem Cells/physiology , Flow Cytometry/methods , Biomarkers/blood , Cells, Cultured , Blotting, Western , Cyclin-Dependent Kinase Inhibitor p16 , Cyclin-Dependent Kinase Inhibitor p21
7.
Braz. arch. biol. technol ; 63: e20190003, 2020. tab, graf
Article in English | LILACS | ID: biblio-1132227

ABSTRACT

Abstract Autologous fibrin matrices derived from the Leukocyte and Platelet Rich Plasma (L-PRP) and Leukocyte and Platelet Rich Fibrin (L-PRF) techniques present great potential to act as a bioactive scaffold in regenerative medicine, contributing to the maintenance of cell viability, proliferation stimulus and differentiation. In contrast, there are few studies that characterize the bioactive potential of these fibrin scaffolds by considering the process of production. The objective of this work was to characterize the intrinsic potential of maintaining cell viability of different fibrin scaffolds containing platelets and leukocytes. In order to achieve that, blood samples from a volunteer were collected and processed to obtain fibrin clots using the suggested techniques. To characterize the potential for in vitro viability, mesenchymal stem cells from human infrapatellar fat were used. The scaffolds were cellularized (1x105 cells/scaffolds) and maintained for 5 and 10 days under culture conditions with Dulbecco's Modified Eagle Medium, without addition of fetal bovine serum, and subsequently subjected to analyses by Fourrier transform infra-red spectroscopy, circular dichroism and fluorescence microscopy. The results demonstrated distinct intrinsic potential viability between the scaffolds, and L-PRP was responsible for promoting higher levels of viability in both periods of analysis. No viable cells were identified in the fibrin matrix used as controls. These results allow us to conclude that both fibrin substrates have presented intrinsic potential for maintaining cell viability, with superior potential exhibited by L-PRP scaffold, and represent promising alternatives for use as bioactive supports in musculoskeletal regenerative medicine.


Subject(s)
Humans , Male , Adult , Adipose Tissue/cytology , Tissue Engineering/methods , Platelet-Rich Plasma/cytology , Mesenchymal Stem Cells/physiology , Platelet-Rich Fibrin/cytology , Cell Survival , Spectroscopy, Fourier Transform Infrared , Tissue Scaffolds , Flow Cytometry
8.
Braz. oral res. (Online) ; 33: e079, 2019. graf
Article in English | LILACS | ID: biblio-1019604

ABSTRACT

Abstract Cell therapy associated with guided bone regeneration (GBR) can be used to treat bone defects under challenging conditions such as osteoporosis. This study aimed to evaluate the effect of mesenchymal stem cells (MSCs) in combination with a poly(vinylidene-trifluoroethylene)/barium titanate (PVDF-TrFE/BT) membrane on bone repair in osteoporotic rats. Osteoporosis was induced in female rats by bilateral removal of the ovaries (OVX) or sham surgery (SHAM), and the osteoporotic condition was characterized after 5 months by microtomographic and morphometric analyses. Calvarial defects were created in osteoporotic rats that immediately received the PVDF-TrFE/BT membrane. After 2 weeks, bone marrow-derived MSCs from healthy rats, characterized by the expression of surface markers using flow cytometry, or phosphate-buffered saline (PBS) (Control) were injected into the defects and bone formation was evaluated 4 weeks post-injection by microtomographic, morphometric, and histological analyses. A reduction in the amount of bone tissue in the femurs of OVX compared with SHAM rats confirmed the osteoporotic condition of the experimental model. More bone formation was observed when the defects were injected with MSCs compared to that with PBS. The modification that we are proposing in this study for the classical GBR approach where cells are locally injected after a membrane implantation may be a promising therapeutic strategy to increase bone formation under osteoporotic condition.


Subject(s)
Animals , Female , Polyvinyls/pharmacology , Titanium/pharmacology , Barium Compounds/pharmacology , Guided Tissue Regeneration/methods , Mesenchymal Stem Cells/physiology , Osteogenesis/drug effects , Osteoporosis/physiopathology , Osteoporosis/therapy , Polyvinyls/chemistry , Time Factors , Titanium/chemistry , Bone Regeneration/drug effects , Bone Regeneration/physiology , Ovariectomy , Random Allocation , Bone Density , Reproducibility of Results , Treatment Outcome , Rats, Wistar , Barium Compounds/chemistry , Imaging, Three-Dimensional , Mesenchymal Stem Cells/chemistry , Flow Cytometry
9.
Electron. j. biotechnol ; 34: 59-66, july. 2018. graf, tab, ilus
Article in English | LILACS | ID: biblio-1047365

ABSTRACT

Background: The use of novel materials as an artificial extracellular matrix for stem cell growth is a current strategy of increasing interest for regenerative medicine. Here, we prepare thermal-remolded membrane scaffolds from poly(3-hydroxybutyrate) grafted with 2-amino-ethyl methacrylate hydrochloride. However, it is unclear whether these membranes are useful for tissue engineering. Results: The mechanical properties, tribology, and morphology of the dense membranes were assessed. The results show that tensile strain at break and roughness of the compressed membrane decrease with increasing graft degree. Moreover, graft copolymer membranes showed lower resistance to scratching, greater degree of swelling and higher brittleness than un-grafted P(3HB) films. Thus, it effectively supports the growth of dermal fibroblast, as demonstrated by epifluorescence microscopy. Conclusions: It is concluded that the developed membrane can be properly used in is the restoration of skin tissue. How to cite: González-Torres M, Sánchez-Sánchez R, Solís-Rosales SG, et al. Poly(3-hydroxybutyrate) graft copolymer dense membranes for human mesenchymal stem cell growth.


Subject(s)
Mesenchymal Stem Cells/physiology , Membranes, Artificial , Temperature , Regenerative Medicine , Growth
10.
Braz. oral res. (Online) ; 32: e83, 2018. tab, graf
Article in English | LILACS | ID: biblio-974444

ABSTRACT

Abstract Distraction osteogenesis (DO) relies on the recruitment and proliferation of mesenchymal stem cells (MSC) to the target site, where they differentiate into osteoblasts to promote bone formation. Nevertheless, MSC recruitment appears to be slow and limits bone formation in DO defects. Thus, this systematic review aims to evaluate the ability of locally applied MSC to enhance bone formation in DO preclinical models. Databases were searched for quantitative pre-clinical controlled studies that evaluated the effect of local administration of MSC on DO bone formation. Eligible studies were identified and data regarding study characteristics, outcome measures and quality were extracted. Nine studies met the inclusion criteria. Autogenous and xenogenous MSC were used to promote DO bone formation. These included bone marrow-derived MSC, adipose tissue-derived MSC and MSC derived from human exfoliated deciduous teeth. Meta-analysis was not possible due to heterogeneities in study designs. Local MSC implantation was not associated with adverse effects. In 4 out of the 5 studies, locally delivered undifferentiated bone-marrow MSC had a positive effect on DO bone formation. Few studies evaluated the therapeutic effects of MSC from other sources. The adjunct use of biologically active molecules or forced expression of key genes involved in osteogenesis further boosted the ability of bone-marrow MSC to promote DO bone formation. While risk of bias and heterogeneity limited the strength of this systematic review, our results suggest that the use of MSC is safe and may provide beneficial effects on DO bone formation.


Subject(s)
Animals , Osteogenesis/physiology , Osteogenesis, Distraction/methods , Models, Animal , Mesenchymal Stem Cell Transplantation/methods , Mesenchymal Stem Cells/physiology , Bone Regeneration/physiology , Bias , Reproducibility of Results , Risk Factors , Treatment Outcome
11.
Journal of Peking University(Health Sciences) ; (6): 962-967, 2018.
Article in Chinese | WPRIM | ID: wpr-941730

ABSTRACT

OBJECTIVE@#To study the effects of disintegrin and metalloproteinase (ADAM) 9, 15 and 17 on the osteogenic differentiation of bone marrow mesenchymal stem cells (BMMSCs).@*METHODS@#BMMSCs of ADAM9, ADAM15, ADAM17 conditional knockout mice and wild type mice (WT) were induced and cultured. Alkaline phosphatase (ALP) activity was measured by colorimetry, early osteogenic transcription factors Runx and Osterix were detected by Real-time PCR, and mineral formation was analyzed by alizarin red staining.@*RESULTS@#ALP activity was lower in ADAM9 group (8.08±0.34), ADAM15 group (6.46±3.40), ADAM17 group (9.30±2.30) than that in WT group (9.44±2.50), but there was no significant difference (P>0.05). Stimulated with bone morphogenetic protein 2(BMP2),there was significant difference (P<0.05) between ADAM9 group (14.22±3.25), ADAM15 group (10.14±2.40) and WT group (20.89±3.40), and ADAM 17 group (23.56±2.50) was higher than WT group (20.89±3.40), but no significant difference (P>0.05). Similarly, cultured by osteogenic induction medium (OST), compared with WT group (12.97±1.30), ADAM9 group (9.63±1.00) and ADAM15 group (7.75±1.30) were lower, ADAM17 group (20.09±1.68) was higher, and the difference was statistically significant (P<0.05). Using stimulated culture by BMP2 and OST combined, ADAM9 group (15.75±1.30), ADAM 15 group (12.43±1.30) were less than WT group (26.15 ±1.50), while ADAM17 group (29.55±2.10) was higher than WT group were statistically significant (P<0.05). The expression of Runx2 in ADAM9 group (2.02±0.24), ADAM15 group (3.09±0.19), ADAM17 group (3.89±0.91) had no significant difference compared with WT (2.02±0.21) group (P>0.05). ADAM9 group stimulated by BMP2 (7.00±0.23), ADAM15 group (6.04±0.23) were lower than WT group (12.6±0.23), ADAM17 group (18.52±1.39) was higher than WT group (12.6±0.23), and the difference was statistically significant (P<0.05). In non-stimulating culture, there was no significant difference in Osterix expression between ADAM9 group (9.60±3.87), ADAM17 group (12.40±3.00) and WT group (10.9±1.10, P>0.05), but in ADAM15 group (6.50±1.51) it was slightly lower than that in WT group (P<0.05). After BMP2 stimulation, ADAM9 group (39.20±3.23) and ADAM15 group (20.50±4.80) were less than WT group (60.30±5.93), while ADAM17 group (80.20±3.30) was higher than WT group (P<0.05). Alizarin red staining showed no obvious orange-red mass in the non-induction group. Local calcified nodules could be seen in the BMP2, OST, OST + BMP2 induction culture conditions in all the experimental groups, but there was no significant difference in quantitative analysis (P>0.05).@*CONCLUSION@#ADAM9, 15, 17 took part in the osteogenic differentiation of BMMSCs, and provided new targets for its regulation.


Subject(s)
Animals , Mice , ADAM Proteins/physiology , Cell Differentiation , Cells, Cultured , Integrins , Mesenchymal Stem Cells/physiology , Mice, Knockout , Osteogenesis
12.
Rev. bras. ginecol. obstet ; 39(5): 217-223, May 2017. tab, graf
Article in English | LILACS | ID: biblio-898862

ABSTRACT

Abstract Purpose To evaluate the effect of mesenchymal stem cells (MSCs) on fertility in experimental retrocervical endometriosis. Methods A total of 27 New Zealand rabbits were divided into three groups: endometriosis, in which endometrial implants were created; mesenchymal, in which MSCs were applied in addition to the creation of endometrial implants; and control, the group without endometriosis. Fisher's exact test was performed to compare the dichotomous qualitative variables among the groups. The quantitative variables were compared by the nonparametric Mann-Whitney and Kruskal-Wallis tests. The MannWhitney test was used for post-hoc multiple comparison with Boniferroni correction. Results Regarding the beginning of the fertile period, the three groups had medians of 14±12.7, 40±5, and 33±8.9 days respectively (p = 0.005). With regard to fertility (number of pregnancies), the endometriosis and control groups showed a rate of 77.78%, whereas the mesenchymal group showed a rate of 11.20% (p = 0.015). No differences in Keenan's histological classification were observed among the groups (p = 0.730). With regard to the macroscopic appearance of the lesions, the mesenchymal group showed the most pelvic adhesions. Conclusion The use of MSCs in endometriosis negatively contributed to fertility, suggesting the role of these cells in the development of this disease.


Resumo Objetivo Avaliar o efeito das células-tronco mesenquimais sobre a fertilidade na endometriose retrocervical experimental. Métodos Um total de 27 coelhas da raça Nova Zelândia foram divididas em três grupos: endometriose, em que os implantes endometriais foram criados; mesenquimal, em que as células-tronco mesenquimais foram aplicadas complementarmente à criação implantes endometriais; e controle, sem endometriose. O teste exato de Fisher foi realizado para comparar variáveis dicotômicas qualitativas entre os grupos. As variáveis quantitativas foram comparadas pelos testes não paramétricos de MannWhitney e Kruskal-Wallis. O teste de Mann-Whitney foi utilizado para a comparação múltipla pós-hoc com correção de Boniferroni. Resultados em relação ao início do período fértil, os grupos endometriose, mesenquimal e controle tiveram medianas de 14±12,7; 40±5; e 33±8,9 dias, respectivamente (p = 0,005). Sobre a taxa de fertilidade (número de gravidezes), os grupos endometriose e controle mostraram uma taxa de 77,78%, enquanto o grupo mesenquimal mostrou uma taxa de 11,20% (p = 0,015). Não foram observadas diferenças na classificação histológica de Keenan entre os grupos (p = 0,730). No que diz respeito à aparência macroscópica das lesões, o grupo mesenquimal mostrou maiores adesões pélvicas. Conclusão O uso de células-tronco mesenquimais na endometriose contribuiu negativamente para a fertilidade, sugerindo o papel dessas células no desenvolvimento da doença.


Subject(s)
Humans , Animals , Uterine Cervical Diseases/etiology , Endometriosis/etiology , Mesenchymal Stem Cells/physiology , Infertility, Female/etiology , Rabbits , Uterine Cervical Diseases/pathology , Disease Models, Animal , Endometriosis/pathology , Infertility, Female/pathology
13.
Braz. j. med. biol. res ; 50(2): e5988, 2017. graf
Article in English | LILACS | ID: biblio-839254

ABSTRACT

This study was undertaken to clarify the role and mechanism of pyruvate dehydrogenase kinase isoform 2 (PDK2) in chondrogenic differentiation of mesenchymal stem cells (MSCs). MSCs were isolated from femurs and tibias of Sprague-Dawley rats, weighing 300-400 g (5 females and 5 males). Overexpression and knockdown of PDK2 were transfected into MSCs and then cell viability, adhesion and migration were assessed. Additionally, the roles of aberrant PDK2 in chondrogenesis markers SRY-related high mobility group-box 6 (Sox6), type ΙΙ procollagen gene (COL2A1), cartilage oligomeric matrix protein (COMP), aggrecan (AGC1), type ΙX procollagen gene (COL9A2) and collagen type 1 alpha 1 (COL1A1) were measured by quantitative reverse-transcription polymerase chain reaction (qRT-PCR). The expressions of c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (MAPK) and extracellular regulated protein kinase (ERK) were measured. Overexpressing PDK2 promoted cell viability, adhesion and inhibited cell migration in MSCs (all P<0.05). qRT-PCR assay showed a potent increase in the mRNA expressions of all chondrogenesis markers in response to overexpressing PDK2 (P<0.01 or P<0.05). PDK2 overexpression also induced a significant accumulation in mRNA and protein expressions of JNK, p38MAPK and ERK in MSCs compared to the control (P<0.01 or P<0.05). Meanwhile, silencing PDK2 exerted the opposite effects on MSCs. This study shows a preliminary positive role and potential mechanisms of PDK2 in chondrogenic differentiation of MSCs. It lays the theoretical groundwork for uncovering the functions of PDK2 and provides a promising basis for repairing cartilage lesions in osteoarthritis.


Subject(s)
Animals , Male , Female , Rats , Chondrogenesis/physiology , JNK Mitogen-Activated Protein Kinases/physiology , MAP Kinase Signaling System/physiology , Mesenchymal Stem Cells/physiology , Protein Serine-Threonine Kinases/physiology , SOXE Transcription Factors/physiology , Cell Differentiation , Rats, Sprague-Dawley , Transcriptional Activation , Up-Regulation
14.
Bauru; s.n; 2017. 93 p. ilus, tab, graf.
Thesis in Portuguese | LILACS, BBO | ID: biblio-880080

ABSTRACT

Inserido no paradigma da transdisciplinaridade, o presente trabalho foi desenvolvido em etapas, com os seguintes objetivos: a) Construir um dispositivo com base de metal não magnético para ímãs permanentes, visando à geração de um Campo Magnético Estático (CME) ou de um Campo Magnético Compensado (CMC); b) Expor culturas de células mesenquimais a um CME e a um CMC, ou a nenhum campo (controle); c) Analisar a influência destes campos na viabilidade e proliferação celular e nos casos em que houve alteração em pelo menos um destes parâmetros, utilizar a análise proteômica como ferramenta para a compreensão dos mecanismos envolvidos. O dispositivo foi construído utilizando aço inoxidável, capaz de gerar dois tipos de Campos Magnéticos: Compensado (CMC) com intensidade de aproximadamente 0 mT e Estático (CME) com intensidade média de 165 mT. Estes campos foram aplicados a culturas de células mesenquimais de medula óssea de camundongos AJ (MSC/AJ), nos períodos de 0, 24, 48, 72 e 96 h (CMC) e 24 h (CME). Os efeitos sobre a proliferação e a viabilidade foram avaliados por método de contagem manual de células com marcação por azul de tripan. A análise proteômica foi realizada para os experimentos com CMC, com o objetivo de descrever as proteínas envolvidas nas alterações encontradas. A exposição ao CMC tendeu a reduzir a proliferação das células de medula óssea MSC/AJ em relação ao controle em 96 h, porém sem diferença significativa, o que poderia estar relacionado a proteínas que inibem a transcrição, como a Forkhead box protein P2 Foxp2. Este mesmo campo aumentou a viabilidade celular em relação ao baseline para todos os tempos experimentais, o que poderia estar relacionado a proteínas relacionadas à ligação ao Ca+2. Esses mecanismos, entretanto, precisam ser estudados mais profundamente para que possam ser comprovados ou não. Já a exposição ao CME levou a uma tendência à diminuição da proliferação e viabilidade celular em relação ao grupo controle, embora sem diferenças significativas, provavelmente por conta do tamanho amostral e tempo de avaliação (24 h).(AU)


Inserted in the transdisciplinarity paradigm, the present work was developed by steps with the following aims: a) To build a device of non-magnetic metal to hold permanent magnets for the generation of a Static Magnetic Field (SMF) or a Compensated Magnetic Field (CMF); b) To expose mesenchimal cells to the SMF and to CMF or to none of the fields (control); c) To analyze the influence of these fields on cell viability and cell proliferation and in the case where it occurred alteration in at least one of these parameters, to use proteomics as a tool for the comprehension of the involved mechanisms. The device was built in stainless steel, able to generate two kinds of Magnetic Fields: Compesated (CMF) with an intensity of nearly zero mT and Static (SMF) with a mean intensity of 165 mT. These fields were applied to bone marrow mesenchimal cell cultures from AJ mice (MSC/AJ), for 0, 24, 48, 72 and 96 h (CMF) and 24 h (SMF) periods. The effects on the proliferation and viability were assessed by tripan blue dying and manual counting of the cells. Proteomics was done for the experiments with CMF, aiming to describe the involved proteins on found alterations. The exposition to CMF tends to reduce the bone marrow cell proliferation of MSC/AJ in relation to control in 96 h, but with no significant difference, which may be related to proteins that inhibit the transcription, like Forkhead box protein P2 Foxp2. This very field raised the cell viability in relation to the baseline for all the experimental times that could be related to proteins connected to Ca2+ binding. However, these mechanisms need more experiments, so they can be confirmed or not. The exposition to the SMF tends to decrease both cell proliferation and viability in relation to the control group, although with no significant difference, probably because of the sample number and the exposition time (24h).(AU)


Subject(s)
Animals , Male , Mice , Cell Proliferation/physiology , Magnetic Fields , Mesenchymal Stem Cells/physiology , Cell Count , Cell Survival/physiology , Cells, Cultured , Chromatography, Liquid , Mass Spectrometry , Reference Values , Reproducibility of Results , Time Factors
15.
J. appl. oral sci ; 24(4): 376-382, July-Aug. 2016. graf
Article in English | LILACS, BBO | ID: lil-792596

ABSTRACT

ABSTRACT Aging negatively affects bone/titanium implant interactions. Our hypothesis is that the unbalance between osteogenesis and adipogenesis induced by aging may be involved in this phenomenon. Objective We investigated the osteoblast and adipocyte differentiation of mesenchymal stem cells (MSCs) from young and aged rats cultured on Ti. Material and Methods Bone marrow MSCs derived from 1-month and 21-month rats were cultured on Ti discs under osteogenic conditions for periods of up to 21 days and osteoblast and adipocyte markers were evaluated. Results Cell proliferation, alkaline phosphatase (ALP) activity, extracellular matrix mineralization and gene expression of RUNX2, osterix, ALP, bone sialoprotein, osteopontin, and osteocalcin were reduced in cultures of 21-month rats compared with 1-month rats grown on Ti. Gene expression of PPAR-γ , adipocyte protein 2, and resistin and lipid accumulation were increased in cultures of 21-month rats compared with 1-month rats grown on the same conditions. Conclusions These results indicate that the lower osteogenic potential of MSCs derived from aged rats compared with young rats goes along with the higher adipogenic potential in cultures grown on Ti surface. This unbalance between osteoblast and adipocyte differentiation should be considered in dental implant therapy to the elderly population.


Subject(s)
Animals , Female , Rats , Osteoblasts/physiology , Titanium/chemistry , Aging/physiology , Dental Implants , Adipogenesis/physiology , Mesenchymal Stem Cells/physiology , Osteogenesis/physiology , Surface Properties , Gene Expression , Cells, Cultured , Age Factors , Cell Proliferation/physiology , Alkaline Phosphatase/analysis , Real-Time Polymerase Chain Reaction , Lipids/analysis
16.
Rev. Ateneo Argent. Odontol ; 55(1): 69-70, 2016. ilus
Article in Spanish | LILACS | ID: lil-794296

ABSTRACT

La utilización de células indiferenciadas embrionarias y de células diferenciadas inducidas para que se comporten como las anteriores permite dar origen adiferentes tejidos que pueden ser usados en medicina reconstructiva en reemplazo de los deteriorados...


Subject(s)
Humans , Multipotent Stem Cells/physiology , Pluripotent Stem Cells/physiology , Totipotent Stem Cells/physiology , Stem Cells/physiology , Plastic Surgery Procedures/methods , Mesenchymal Stem Cells/physiology , Fetal Stem Cells/physiology , Tissue Engineering/methods
17.
Gut and Liver ; : 166-176, 2016.
Article in English | WPRIM | ID: wpr-25631

ABSTRACT

Mesothelial cells (MCs) cover the surface of visceral organs and the parietal walls of cavities, and they synthesize lubricating fluids to create a slippery surface that facilitates movement between organs without friction. Recent studies have indicated that MCs play active roles in liver development, fibrosis, and regeneration. During liver development, the mesoderm produces MCs that form a single epithelial layer of the mesothelium. MCs exhibit an intermediate phenotype between epithelial cells and mesenchymal cells. Lineage tracing studies have indicated that during liver development, MCs act as mesenchymal progenitor cells that produce hepatic stellate cells, fibroblasts around blood vessels, and smooth muscle cells. Upon liver injury, MCs migrate inward from the liver surface and produce hepatic stellate cells or myofibroblast depending on the etiology, suggesting that MCs are the source of myofibroblasts in capsular fibrosis. Similar to the activation of hepatic stellate cells, transforming growth factor β induces the conversion of MCs into myofibroblasts. Further elucidation of the biological and molecular changes involved in MC activation and fibrogenesis will contribute to the development of novel approaches for the prevention and therapy of liver fibrosis.


Subject(s)
Humans , Epithelial Cells/physiology , Epithelium/metabolism , Hepatic Stellate Cells/physiology , Liver/cytology , Liver Cirrhosis/etiology , Liver Regeneration/physiology , Mesenchymal Stem Cells/physiology , Myofibroblasts/physiology
18.
Int. braz. j. urol ; 41(5): 990-1001, Sept.-Oct. 2015. graf
Article in English | LILACS | ID: lil-767059

ABSTRACT

ABSTRACT Objectives: Diseases of the genitourinary tract can lead to significant damage. Current reconstructive techniques are limited by tissue availability and compatibility. This study aims to assess if the decellularized human glans can be used as a biomaterial for penile reconstruction. Materials and Methods: Samples of the glans matrices were descellularized. We evaluate the presence of collagen type I and III, and elastic fibers. Biocompatibility assays were performed to assess the cytotoxic and non-cytotoxic interactions between the acellular matrix and 3T3 cells. The matrices were seeded with mesenchymal stem cells and were assessed for viability and integration of these cells. Biomechanical tests in native tissue, descellularized matrix and seeded matrix were performed to characterize their biomechanical properties. Results: The tissue architecture of the decellularized matrix of human glans was preserved as well as the maintenance of the biomechanical and biological properties. The analyzes of glans seeded with mesenchymal stem cells revealed the integration of these cells to the matrices, and its viability during two weeks "in vitro". Conclusion: The decellularization process did not alter the biological and biomechanical characteristics of the human glans. When these matrices were seeded they were able to maintain the cells integrity and vitality.


Subject(s)
Animals , Humans , Male , Mice , Biocompatible Materials , Extracellular Matrix/physiology , Mesenchymal Stem Cells/physiology , Penis/cytology , Tissue Scaffolds , Tissue Engineering/methods , /physiology , Biomechanical Phenomena , Cells, Cultured , Collagen Type I/analysis , Collagen Type II/analysis , Materials Testing , Mesenchymal Stem Cells/cytology , Rats, Wistar , Reproducibility of Results , Time Factors
19.
Rev. bras. cir. cardiovasc ; 30(3): 380-385, July-Sept. 2015. tab
Article in English | LILACS | ID: lil-756523

ABSTRACT

AbstractPulmonary hypertension is a devastating and refractory disease and there is no cure for this disease. Recently, microRNAs and mesenchymal stem cells emerged as novel methods to treat pulmonary hypertension. More than 20 kinds of microRNAs may participate in the process of pulmonary hypertension. It seems microRNAs or mesenchymal stem cells can ameliorate some symptoms of pulmonary hypertension in animals and even improve heart and lung function during pulmonary hypertension. Nevertheless, the relationship between mesenchymal stem cells, microRNAs and pulmonary hypertension is not clear. And the mechanisms underlying their function still need to be investigated. In this study we review the recent findings in mesenchymal stem cells - and microRNAs-based pulmonary hypertension treatment, focusing on the potential role of microRNAs regulated mesenchymal stem cells in pulmonary hypertension and the role of exosomes between mesenchymal stem cells and pulmonary hypertension.


ResumoA hipertensão pulmonar é uma doença devastadora e refratária, para a qual não existe cura. Recentemente, microRNAs e células-tronco mesenquimais emergiram como novos métodos para tratar a hipertensão pulmonar. Mais de 20 tipos de microRNAs podem participar no processo de hipertensão pulmonar. Ao que parece, microRNAs ou células-tronco mesenquimais podem atenuar alguns sintomas de hipertensão pulmonar em animais de e até mesmo melhorar a função cardíaca e do pulmão durante a hipertensão pulmonar. No entanto, a relação entre células-tronco mesenquimais, microRNAs e hipertensão pulmonar não é clara. E os mecanismos subjacentes a sua função ainda precisam ser investigados. Neste estudo, revisamos as descobertas recentes no tratamento da hipertensão pulmonar baseado em células-tronco mesenquimais e microRNAs, enfocando o papel potencial dos microRNAs para regular as células-tronco mesenquimais na hipertensão pulmonar e o papel dos exossomos entre células-tronco mesenquimais e hipertensão pulmonar.


Subject(s)
Animals , Humans , Hypertension, Pulmonary/therapy , Mesenchymal Stem Cell Transplantation/methods , Mesenchymal Stem Cells/physiology , MicroRNAs/therapeutic use , Exosomes/physiology , Hypertension, Pulmonary/physiopathology , Lung/physiopathology
20.
Biol. Res ; 48: 1-8, 2015. ilus, graf
Article in English | LILACS | ID: biblio-950833

ABSTRACT

BACKGROUND: Human amnion mesenchymal cells (hAMCs), isolated from the amniotic membrane of human placenta, are a unique population of mesenchymal stem cells. Recent studies demonstrated that hAMCs could inhibit the activities and functions of several immune cells. However, their effect on inflammatory macrophages is largely unknown. This study investigated the effect of hAMCs on expression of inflammatory cytokines and mitogen-activated protein kinases (MAPKs)/NF-kB pathway in human THP-1 macrophages induced by lipopolysaccharide (LPS). RESULTS: The levels of TNF-α and IL-1ß secreted by LPS- stimulated THP-1 cells were increased significantly compared with those in the control group. After co-culture with different numbers of hAMCs, the levels of TNF-α and IL-1ß in LPS-stimulated THP-1 cells were significantly reduced compared with the LPS group. The mRNA expression of TNF-α and IL-1ß were also markedly inhibited. Moreover, treating LPS-stimulated THP-1 cells with hAMCs supernatants could also suppress TNF-α and IL-1ß production in THP-1 cells. Important signaling pathways involved in the production of TNF-α and IL-1ß were affected by hAMCs co-culture: hAMCs remarkably suppressed NF-kB activation and down-regulated the phosphorylation of ERK and JNK in LPS- stimulated THP-1 cells. CONCLUSIONS: Human amnion mesenchymal cells inhibited the production of TNF-α and IL-1ß secreted by LPS-stimulated THP-1 cells, partly through the suppression of NF-kB activation and ERK and JNK phosphorylation.


Subject(s)
Humans , Lipopolysaccharides/pharmacology , Tumor Necrosis Factor-alpha/biosynthesis , Interleukin-1beta/biosynthesis , Mesenchymal Stem Cells/physiology , Amnion/cytology , Macrophages/metabolism , Tumor Necrosis Factor-alpha/drug effects , MAP Kinase Signaling System/drug effects , Interleukin-1beta/drug effects
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