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1.
Cells ; 10(6)2021 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-34203839

RESUMO

The bone marrow (BM) is key to protective immunological memory because it harbors a major fraction of the body's plasma cells, memory CD4+ and memory CD8+ T-cells. Despite its paramount significance for the human immune system, many aspects of how the BM enables decade-long immunity against pathogens are still poorly understood. In this review, we discuss the relationship between BM survival niches and long-lasting humoral immunity, how intrinsic and extrinsic factors define memory cell longevity and show that the BM is also capable of adopting many responsibilities of a secondary lymphoid organ. Additionally, with more and more data on the differentiation and maintenance of memory T-cells and plasma cells upon vaccination in humans being reported, we discuss what factors determine the establishment of long-lasting immunological memory in the BM and what we can learn for vaccination technologies and antigen design. Finally, using these insights, we touch on how this holistic understanding of the BM is necessary for the development of modern and efficient vaccines against the pandemic SARS-CoV-2.


Assuntos
Imunidade Adaptativa/fisiologia , Medula Óssea/fisiologia , Plasmócitos/citologia , Linfócitos T/citologia , Vacinologia , Células da Medula Óssea/citologia , Células da Medula Óssea/fisiologia , COVID-19/imunologia , COVID-19/prevenção & controle , Vacinas contra COVID-19/imunologia , Humanos , Imunidade Celular/fisiologia , Memória Imunológica/fisiologia , SARS-CoV-2/imunologia , Linfócitos T/imunologia , Vacinologia/métodos , Vacinologia/tendências
2.
Int J Mol Sci ; 22(12)2021 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-34203758

RESUMO

Synovial fluid contains cytokines, growth factors and resident mesenchymal stem cells (MSCs). The present study aimed to (1) determine the effects of autologous and allogeneic synovial fluid on viability, proliferation and chondrogenesis of equine bone marrow MSCs (BMMSCs) and (2) compare the immunomodulatory properties of equine synovial fluid MSCs (SFMSCs) and BMMSCs after stimulation with interferon gamma (INF-γ). To meet the first aim of the study, the proliferation and viability of MSCs were evaluated by MTS and calcein AM staining assays. To induce chondrogenesis, MSCs were cultured in a medium containing TGF-ß1 or different concentrations of synovial fluid. To meet the second aim, SFMSCs and BMMSCs were stimulated with IFN-γ. The concentration of indoleamine-2,3-dioxygenase (IDO) and nitric oxide (NO) were examined. Our results show that MSCs cultured in autologous or allogeneic synovial fluid could maintain proliferation and viability activities. Synovial fluid affected chondrocyte differentiation significantly, as indicated by increased glycosaminoglycan contents, compared to the chondrogenic medium containing 5 ng/mL TGF-ß1. After culturing with IFN-γ, the conditioned media of both BMMSCs and SFMSCs showed increased concentrations of IDO, but not NO. Stimulating MSCs with synovial fluid or IFN-γ could enhance chondrogenesis and anti-inflammatory activity, respectively, suggesting that the joint environment is suitable for chondrogenesis.


Assuntos
Condrogênese/efeitos dos fármacos , Imunomodulação/efeitos dos fármacos , Interferon gama/farmacologia , Células-Tronco Mesenquimais/imunologia , Líquido Sinovial/metabolismo , Animais , Células da Medula Óssea/citologia , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Ensaio de Unidades Formadoras de Colônias , Cavalos , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Células-Tronco Mesenquimais/enzimologia , Óxido Nítrico/metabolismo
3.
Cells ; 10(6)2021 06 15.
Artigo em Inglês | MEDLINE | ID: covidwho-1273393

RESUMO

The bone marrow (BM) is key to protective immunological memory because it harbors a major fraction of the body's plasma cells, memory CD4+ and memory CD8+ T-cells. Despite its paramount significance for the human immune system, many aspects of how the BM enables decade-long immunity against pathogens are still poorly understood. In this review, we discuss the relationship between BM survival niches and long-lasting humoral immunity, how intrinsic and extrinsic factors define memory cell longevity and show that the BM is also capable of adopting many responsibilities of a secondary lymphoid organ. Additionally, with more and more data on the differentiation and maintenance of memory T-cells and plasma cells upon vaccination in humans being reported, we discuss what factors determine the establishment of long-lasting immunological memory in the BM and what we can learn for vaccination technologies and antigen design. Finally, using these insights, we touch on how this holistic understanding of the BM is necessary for the development of modern and efficient vaccines against the pandemic SARS-CoV-2.


Assuntos
Imunidade Adaptativa/fisiologia , Medula Óssea/fisiologia , Plasmócitos/citologia , Linfócitos T/citologia , Vacinologia , Células da Medula Óssea/citologia , Células da Medula Óssea/fisiologia , COVID-19/imunologia , COVID-19/prevenção & controle , Vacinas contra COVID-19/imunologia , Humanos , Imunidade Celular/fisiologia , Memória Imunológica/fisiologia , SARS-CoV-2/imunologia , Linfócitos T/imunologia , Vacinologia/métodos , Vacinologia/tendências
4.
Molecules ; 26(11)2021 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-34205205

RESUMO

Total body irradiation is a standard procedure of bone marrow transplantation (BMT) which causes a rapid increase in reactive oxygen species (ROS) in the bone marrow microenvironment during BMT. The increase in ROS reduces the engraftment ability of donor cells, thereby affecting the bone marrow recovery of recipients after BMT. In the early weeks following transplantation, recipients are at high risk of severe infection due to weakened hematopoiesis. Thus, it is imperative to improve engraftment capacity and accelerate bone marrow recovery in BMT recipients. In this study, we constructed recombinant copper/zinc superoxide dismutase 1 (SOD1) fused with the cell-penetrating peptide (CPP), the trans-activator of transcription (Tat), and showed that this fusion protein has penetrating ability and antioxidant activity in both RAW264.7 cells and bone marrow cells in vitro. Furthermore, irradiated mice transplanted with SOD1-Tat-treated total bone marrow donor cells showed an increase in total bone marrow engraftment capacity two weeks after transplantation. This study explored an innovative method for enhancing engraftment efficiency and highlights the potential of CPP-SOD1 in ROS manipulation during BMT.


Assuntos
Antioxidantes/farmacologia , Células da Medula Óssea/citologia , Peptídeos Penetradores de Células/genética , Produtos do Gene tat/genética , Proteínas Recombinantes de Fusão/farmacologia , Superóxido Dismutase-1/genética , Animais , Antioxidantes/metabolismo , Células da Medula Óssea/efeitos dos fármacos , Transplante de Medula Óssea , Peptídeos Penetradores de Células/metabolismo , Células Cultivadas , Produtos do Gene tat/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Espécies Reativas de Oxigênio , Proteínas Recombinantes de Fusão/metabolismo , Superóxido Dismutase-1/metabolismo , Irradiação Corporal Total
5.
Int J Mol Sci ; 22(11)2021 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-34072546

RESUMO

Non-targeted effects (NTE) of ionizing radiation may initiate myeloid neoplasms (MN). Here, protein mediators (I) in irradiated human mesenchymal stromal cells (MSC) as the NTE source, (II) in MSC conditioned supernatant and (III) in human bone marrow CD34+ cells undergoing genotoxic NTE were investigated. Healthy sublethal irradiated MSC showed significantly increased levels of reactive oxygen species. These cells responded by increasing intracellular abundance of proteins involved in proteasomal degradation, protein translation, cytoskeleton dynamics, nucleocytoplasmic shuttling, and those with antioxidant activity. Among the increased proteins were THY1 and GNA11/14, which are signaling proteins with hitherto unknown functions in the radiation response and NTE. In the corresponding MSC conditioned medium, the three chaperones GRP78, CALR, and PDIA3 were increased. Together with GPI, these were the only four altered proteins, which were associated with the observed genotoxic NTE. Healthy CD34+ cells cultured in MSC conditioned medium suffered from more than a six-fold increase in γH2AX focal staining, indicative for DNA double-strand breaks, as well as numerical and structural chromosomal aberrations within three days. At this stage, five proteins were altered, among them IQGAP1, HMGB1, and PA2G4, which are involved in malign development. In summary, our data provide novel insights into three sequential steps of genotoxic signaling from irradiated MSC to CD34+ cells, implicating that induced NTE might initiate the development of MN.


Assuntos
Células da Medula Óssea/metabolismo , Diferenciação Celular , Dano ao DNA , Células-Tronco Mesenquimais/metabolismo , Proteoma , Transdução de Sinais , Idoso , Antígenos CD34/metabolismo , Biomarcadores , Células da Medula Óssea/citologia , Diferenciação Celular/genética , Diferenciação Celular/efeitos da radiação , Sobrevivência Celular/genética , Instabilidade Cromossômica , Meios de Cultivo Condicionados/metabolismo , Feminino , Histonas/metabolismo , Humanos , Masculino , Células-Tronco Mesenquimais/citologia , Modelos Biológicos , Proteômica/métodos , Radiação Ionizante , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos da radiação
6.
Int J Mol Sci ; 22(11)2021 May 26.
Artigo em Inglês | MEDLINE | ID: mdl-34073480

RESUMO

Mesenchymal stem cells (MSCs) can differentiate into osteoblasts, and therapeutic targeting of these cells is considered both for malignant and non-malignant diseases. We analyzed global proteomic profiles for osteoblasts derived from ten and MSCs from six healthy individuals, and we quantified 5465 proteins for the osteoblasts and 5420 proteins for the MSCs. There was a large overlap in the profiles for the two cell types; 156 proteins were quantified only in osteoblasts and 111 proteins only for the MSCs. The osteoblast-specific proteins included several extracellular matrix proteins and a network including 27 proteins that influence intracellular signaling (Wnt/Notch/Bone morphogenic protein pathways) and bone mineralization. The osteoblasts and MSCs showed only minor age- and sex-dependent proteomic differences. Finally, the osteoblast and MSC proteomic profiles were altered by ex vivo culture in serum-free media. We conclude that although the proteomic profiles of osteoblasts and MSCs show many similarities, we identified several osteoblast-specific extracellular matrix proteins and an osteoblast-specific intracellular signaling network. Therapeutic targeting of these proteins will possibly have minor effects on MSCs. Furthermore, the use of ex vivo cultured osteoblasts/MSCs in clinical medicine will require careful standardization of the ex vivo handling of the cells.


Assuntos
Células da Medula Óssea/metabolismo , Regulação da Expressão Gênica , Células-Tronco Mesenquimais/metabolismo , Osteoblastos/metabolismo , Proteômica , Via de Sinalização Wnt , Idoso , Células da Medula Óssea/citologia , Feminino , Humanos , Masculino , Células-Tronco Mesenquimais/citologia , Pessoa de Meia-Idade , Osteoblastos/citologia
7.
Cell Prolif ; 54(7): e13058, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34053135

RESUMO

OBJECTIVES: To clarify the possible role and mechanism of Cathepsin K (CTSK) in alveolar bone regeneration mediated by jaw bone marrow mesenchymal stem cells (JBMMSC). MATERIALS AND METHODS: Tooth extraction models of Ctsk knockout mice (Ctsk-/- ) and their wildtype (WT) littermates were used to investigate the effect of CTSK on alveolar bone regeneration. The influences of deletion or inhibition of CTSK by odanacatib (ODN) on proliferation and osteogenic differentiation of JBMMSC were assessed by CCK-8, Western blot and alizarin red staining. To explore the differently expressed genes, RNA from WT and Ctsk-/- JBMMSC was sent to RNA-seq. ECAR, glucose consumption and lactate production were measured to identify the effect of Ctsk deficiency or inhibition on glycolysis. At last, we explored whether Ctsk deficiency or inhibition promoted JBMMSC proliferation and osteogenic differentiation through glycolysis. RESULTS: We found out that Ctsk knockout could promote alveolar bone regeneration in vivo. In vitro, we confirmed that both Ctsk knockout and inhibition by ODN could promote proliferation of JBMMSC, up-regulate expression of Runx2 and ALP, and enhance matrix mineralization. RNA-seq results showed that coding genes of key enzymes in glycolysis were significantly up-regulated in Ctsk-/- JBMMSC, and Ctsk deficiency or inhibition could promote glycolysis in JBMMSC. After blocking glycolysis by 3PO, the effect of Ctsk deficiency or inhibition on JBMMSC's regeneration was blocked subsequently. CONCLUSIONS: Our findings revealed that Ctsk knockout or inhibition could promote alveolar bone regeneration by enhancing JBMMSC regeneration via glycolysis. These results shed new lights on the regulatory mechanism of CTSK on bone regeneration.


Assuntos
Regeneração Óssea , Catepsina K/genética , Diferenciação Celular , Proliferação de Células , Células-Tronco Mesenquimais/metabolismo , Animais , Células da Medula Óssea/citologia , Catepsina K/deficiência , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/metabolismo , Glucose/metabolismo , Glicólise , Arcada Osseodentária/citologia , Células-Tronco Mesenquimais/citologia , Camundongos , Camundongos Knockout , Osteoblastos/citologia , Osteoblastos/metabolismo , Osteogênese
8.
Nat Commun ; 12(1): 2665, 2021 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-33976125

RESUMO

With age, hematopoietic stem cells (HSC) undergo changes in function, including reduced regenerative potential and loss of quiescence, which is accompanied by a significant expansion of the stem cell pool that can lead to haematological disorders. Elevated metabolic activity has been implicated in driving the HSC ageing phenotype. Here we show that nicotinamide riboside (NR), a form of vitamin B3, restores youthful metabolic capacity by modifying mitochondrial function in multiple ways including reduced expression of nuclear encoded metabolic pathway genes, damping of mitochondrial stress and a decrease in mitochondrial mass and network-size. Metabolic restoration is dependent on continuous NR supplementation and accompanied by a shift of the aged transcriptome towards the young HSC state, more youthful bone marrow cellular composition and an improved regenerative capacity in a transplant setting. Consequently, NR administration could support healthy ageing by re-establishing a more youthful hematopoietic system.


Assuntos
Envelhecimento , Células-Tronco Hematopoéticas/efeitos dos fármacos , NAD/metabolismo , Niacinamida/análogos & derivados , Compostos de Piridínio/farmacologia , Fatores Etários , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/metabolismo , Células Cultivadas , Perfilação da Expressão Gênica/métodos , Regulação da Expressão Gênica/efeitos dos fármacos , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Modelos Biológicos , Niacinamida/farmacologia , Fosforilação Oxidativa/efeitos dos fármacos
9.
Nat Commun ; 12(1): 2856, 2021 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-34001893

RESUMO

Neutrophils are implicated in multiple homeostatic and pathological processes, but whether functional diversity requires discrete neutrophil subsets is not known. Here, we apply single-cell RNA sequencing to neutrophils from normal and inflamed mouse tissues. Whereas conventional clustering yields multiple alternative organizational structures, diffusion mapping plus RNA velocity discloses a single developmental spectrum, ordered chronologically. Termed here neutrotime, this spectrum extends from immature pre-neutrophils, largely in bone marrow, to mature neutrophils predominantly in blood and spleen. The sharpest increments in neutrotime occur during the transitions from pre-neutrophils to immature neutrophils and from mature marrow neutrophils to those in blood. Human neutrophils exhibit a similar transcriptomic pattern. Neutrophils migrating into inflamed mouse lung, peritoneum and joint maintain the core mature neutrotime signature together with new transcriptional activity that varies with site and stimulus. Together, these data identify a single developmental spectrum as the dominant organizational theme of neutrophil heterogeneity.


Assuntos
Neutrófilos/metabolismo , Análise de Sequência de RNA/métodos , Análise de Célula Única/métodos , Transcriptoma/genética , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Células Cultivadas , Feminino , Ontologia Genética , Humanos , Masculino , Camundongos Endogâmicos C57BL , Neutrófilos/citologia , Peritonite/genética , Peritonite/patologia , Pneumonia/genética , Pneumonia/patologia , Baço/citologia , Baço/metabolismo
10.
Biochem Biophys Res Commun ; 556: 171-178, 2021 06 04.
Artigo em Inglês | MEDLINE | ID: mdl-33839412

RESUMO

It is well known that osteoporosis is a significant chronic disease with the increase of the aging population. Here, we report that expression of G protein-coupled receptor 35 (GPR35) in bone marrow mesenchymal stem cells (BMSCs) is suppressed in diagnosed osteoporosis patients and osteoporotic mice. The expression of GPR35 on BMSCs is enhanced during osteogenic differentiation. GPR35 knockout suppresses the proliferation and osteogenesis of BMSCs and deteriorates bone mass in both sham-treated and ovariectomized mice. Moreover, GPR35 deficiency reduces ß-catenin activity in BMSCs. In contrast, the overexpression of GPR35 contributes to these processes in BMSCs. Finally, using zaprinast, a synthetic GPR35 agonist, we show that zaprinast rescues OVX-induced bone loss and promotes bone generation in mice. Thus, GPR35 may as a new target and its agonist zaprinast may serve as a novel treatment for osteoporosis.


Assuntos
Glicogênio Sintase Quinase 3 beta/metabolismo , Osteogênese , Receptores Acoplados a Proteínas G/metabolismo , Via de Sinalização Wnt , beta Catenina/metabolismo , Animais , Densidade Óssea/efeitos dos fármacos , Densidade Óssea/genética , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Osso e Ossos/efeitos dos fármacos , Osso e Ossos/metabolismo , Proliferação de Células , Feminino , Humanos , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Tamanho do Órgão/efeitos dos fármacos , Tamanho do Órgão/genética , Osteoporose Pós-Menopausa/tratamento farmacológico , Osteoporose Pós-Menopausa/prevenção & controle , Ovariectomia , Purinonas/farmacologia , Purinonas/uso terapêutico , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/deficiência , Receptores Acoplados a Proteínas G/genética
11.
Life Sci ; 277: 119520, 2021 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-33887345

RESUMO

AIMS: Mouse bone marrow mesenchymal stem cells (BMSCs) are pluripotent cells with self-renewal and differentiation abilities. Since the effects of senescent BMSCs on C2C12 cells are not fully clear, the present study aimed to elucidate these effects. MAIN METHODS: Senescence-associated ß-galactosidase staining and western blotting were performed to confirm the senescence of BMSCs. Immunofluorescence and western blotting were used to assess myoblast differentiation in each group. The role of the AKT/P70 signaling pathway and forkhead box O3 (FOXO3) nuclear translocation was explored by western blotting. BMSC-derived exosomes were injected into the tibialis anterior of mice, and RT-qPCR was used to assess the role of exosomes in promoting muscle differentiation. KEY FINDINGS: Conditioned medium (CM) from early-senescent BMSCs promoted myogenic differentiation in vitro, which was detected as enhanced expression of myosin heavy chain (MHC), myogenin (MYOG), and myogenic differentiation 1 (MyoD). The AKT signaling pathway was found to be regulated by CM, which inhibited FOXO3 nuclear translocation. RT-qPCR analysis results showed that MHC, MyoD, and MYOG mRNA expression increased in the tibialis anterior of mice after exosome injection. SIGNIFICANCE: The present study demonstrated that early-senescent BMSCs accelerated C2C12 cell myogenic differentiation, and the transcription factor, FOXO3, was the target of senescent cells. Collectively, our results suggest that the AKT/P70 signaling pathway mediates the effect of BMSCs on neighboring cells.


Assuntos
Diferenciação Celular/fisiologia , Proteína Forkhead Box O3/metabolismo , Células-Tronco Mesenquimais/metabolismo , Transporte Ativo do Núcleo Celular , Animais , Células da Medula Óssea/citologia , Linhagem Celular , Núcleo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Senescência Celular/efeitos dos fármacos , Células-Tronco Mesenquimais/fisiologia , Camundongos , Desenvolvimento Muscular/fisiologia , Músculo Esquelético/metabolismo , Mioblastos/metabolismo , Miogenina/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo
12.
Biochem Biophys Res Commun ; 551: 7-13, 2021 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-33713981

RESUMO

Both the Nuclear factor-erythroid 2 p45-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) antioxidant pathway and Nucleotide-binding oligomerization domain (NOD)-like receptor protein 3 (NLRP3) pathway are considered essential for the development of acute lung injury (ALI)/ARDS induced by sepsis. Our aim was to study the role of Nrf2/HO-1 pathway on activation of the NLRP3 in the protective effect of marrow mesenchymal stem cells (BMSCs) on LPS-induced ALI. We found that BMSCs ameliorated ALI as evidenced by 1) decreased histopathological injury, wet/dry ratio, and protein permeability index in lung; 2) decreased reactive oxygen species (ROS), malondialdehyde (MDA), and protein carbonyl content and restored the activity of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT) in lung tissue; 3) reduced LPS-induced increase in inflammatory cell count and promotion of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, and IL-6 levels in bronchoalveolar lavage fluid (BALF); 4) improvement in the four-day survival rate of animals; and 5) enhanced expression of Nrf2 and HO-1 and decreased expression of NOD-like receptor protein 3(NLRP3) and caspase-1 (p20) in lung tissue. Of note, Nrf2 transcription factor inhibitor brusatol and HO-1 inhibitor tin protoporphyrin IX (SnppIX) reversed BMSCs induced down-expression of NLRP3 and caspase-1 (p20), and inhibited the protective effects of BMSCs. These findings demonstrated that the Nrf2-mediated HO-1 signaling pathway plays a critical role in the protective effects of BMSCs on LPS-induced ALI. BMSCs may play an anti-inflammatory effect partly through the Nrf2/HO-1-dependent NLRP3 pathway.


Assuntos
Lesão Pulmonar Aguda/metabolismo , Lesão Pulmonar Aguda/prevenção & controle , Células da Medula Óssea/citologia , Endotoxinas/efeitos adversos , Heme Oxigenase (Desciclizante)/metabolismo , Células-Tronco Mesenquimais/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Lesão Pulmonar Aguda/induzido quimicamente , Animais , Antioxidantes/metabolismo , Permeabilidade Capilar/efeitos dos fármacos , Regulação da Expressão Gênica , Heme Oxigenase (Desciclizante)/antagonistas & inibidores , Inflamassomos/efeitos dos fármacos , Inflamassomos/metabolismo , Inflamação/induzido quimicamente , Inflamação/metabolismo , Pulmão/irrigação sanguínea , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Pulmão/patologia , Masculino , Células-Tronco Mesenquimais/citologia , Fator 2 Relacionado a NF-E2/antagonistas & inibidores , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Transdução de Sinais , Taxa de Sobrevida
13.
Int Immunopharmacol ; 95: 107567, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33756225

RESUMO

Coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a global pandemic. However, the mechanism of tissue tropism of SARS-CoV-2 remains unclear. Here, recombinant receptor-binding subdomain 1 of spike protein of SARS-CoV-2 (RBD-SD1) was used as a probe to investigate the potential tropism of SARS-CoV-2 in thirty-three types of normal human tissues. RBD-SD1 probe was observed to interact with cells in reported SARS-CoV-2 infected organs. Interestingly, the RBD-SD1 probe strongly interacted with bone marrow cells in an angiotensin-converting enzyme 2 (ACE2)-independent manner. In addition, SARS-CoV-2 induced the ACE2 mRNA expression in human primary bone marrow cells, suggesting human bone marrow cells may be sensitive to SARS-CoV-2 infection. Therefore, human bone marrow cells could be strongly infected by SARS-CoV-2, which may play an important role in the pathogenesis of COVID-19. These findings provide a deeper understanding of SARS-CoV-2 infection routes, thus contributing to the treatment of COVID-19.


Assuntos
SARS-CoV-2/química , SARS-CoV-2/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Enzima de Conversão de Angiotensina 2/biossíntese , Enzima de Conversão de Angiotensina 2/genética , Células da Medula Óssea/citologia , Células da Medula Óssea/metabolismo , Humanos , Pulmão/citologia , Pulmão/metabolismo , Cultura Primária de Células , Ligação Proteica , Domínios Proteicos , Regulação para Cima
14.
Biomed Res Int ; 2021: 6666836, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33553429

RESUMO

Osteoporosis (OP) is a metabolic disease characterized by decreased bone mass and increased risk of fragility fractures, which significantly reduces the quality of life. Stem cell-based therapies, especially using bone marrow mesenchymal stem cells (BMSCs), are a promising strategy for treating OP. Nevertheless, the survival and differentiation rates of the transplanted BMSCs are low, which limits their therapeutic efficiency. Icariin (ICA) is a traditional Chinese medicine formulation that is prescribed for tonifying the kidneys. It also promotes the proliferation and osteogenic differentiation of BMSCs, although the specific mechanism remains unclear. Based on our previous research, we hypothesized that ICA promotes bone formation via the sclerostin/Wnt/ß-catenin signaling pathway. We isolated rat BMSCs and transfected them with sclerostin gene (SOST) overexpressing or knockdown constructs and assessed osteogenic induction in the presence or absence of ICA. Sclerostin significantly inhibited BMSC proliferation and osteogenic differentiation, whereas the presence of ICA not only increased the number of viable BMSCs but also enhanced ALP activity and formation of calcium nodules during osteogenic induction. In addition, the osteogenic genes including Runx2, ß-catenin, and c-myc as well as antioxidant factors (Prdx1, Cata, and Nqo1) were downregulated by sclerostin and restored by ICA treatment. Mechanistically, ICA exerted these effects by activating the Wnt/ß-catenin pathway. In conclusion, ICA can promote the proliferation and osteogenic differentiation of BMSCs in situ and therefore may enhance the therapeutic efficiency of BMSC transplantation in OP.


Assuntos
Proteínas Morfogenéticas Ósseas/metabolismo , Flavonoides/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Osteogênese/efeitos dos fármacos , Via de Sinalização Wnt/efeitos dos fármacos , Animais , Antioxidantes/metabolismo , Células da Medula Óssea/citologia , Proteínas Morfogenéticas Ósseas/genética , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Medicamentos de Ervas Chinesas/farmacologia , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Marcadores Genéticos/genética , Células-Tronco Mesenquimais/metabolismo , Osteogênese/genética , Osteogênese/fisiologia , Ratos Sprague-Dawley , beta Catenina/metabolismo
15.
Stem Cells Transl Med ; 10(6): 883-894, 2021 06.
Artigo em Inglês | MEDLINE | ID: covidwho-1060671

RESUMO

While mesenchymal stromal cells are an appealing therapeutic option for a range of clinical applications, their potential to induce clotting when used systemically remains a safety concern, particularly in hypercoagulable conditions, such as in patients with severe COVID-19, trauma, or cancers. Here, we tested a novel preclinical approach aimed at improving the safety of mesenchymal stromal cell (MSC) systemic administration by use of a bioreactor. In this system, MSCs are seeded on the exterior of a hollow-fiber filter, sequestering them behind a hemocompatible semipermeable membrane with defined pore-size and permeability to allow for a molecularly defined cross talk between the therapeutic cells and the whole blood environment, including blood cells and signaling molecules. The potential for these bioreactor MSCs to induce clots in coagulable plasma was compared against directly injected "free" MSCs, a model of systemic administration. Our results showed that restricting MSCs exposure to plasma via a bioreactor extends the time necessary for clot formation to occur when compared with "free" MSCs. Measurement of cell surface data indicates the presence of known clot inducing factors, namely tissue factor and phosphatidylserine. Results also showed that recovering cells and flushing the bioreactor prior to use further prolonged clot formation time. Furthermore, application of this technology in two in vivo models did not require additional heparin in fully anticoagulated experimental animals to maintain target activated clotting time levels relative to heparin anticoagulated controls. Taken together the clinical use of bioreactor housed MSCs could offer a novel method to control systemic MSC exposure and prolong clot formation time.


Assuntos
Reatores Biológicos , COVID-19/terapia , Técnicas de Cultura de Células/métodos , Transplante de Células-Tronco Mesenquimais/métodos , Trombose/prevenção & controle , Animais , Anticoagulantes/farmacologia , Testes de Coagulação Sanguínea , Células da Medula Óssea/citologia , Células Cultivadas , Cães , Heparina/farmacologia , Humanos , Masculino , Membranas Artificiais , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Infarto do Miocárdio/patologia , Infarto do Miocárdio/prevenção & controle , SARS-CoV-2 , Suínos
16.
FASEB J ; 35(3): e21413, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33570785

RESUMO

Successful intrauterine hematopoietic cell transplantation (IUT) for congenital hemoglobinopathies is hampered by maternal alloresponsiveness. We investigate these interactions in semi-allogenic murine IUT. E14 fetuses (B6 females × BALB/c males) were each treated with 5E+6 maternal (B6) or paternal (BALB/c) bone marrow cells and serially monitored for chimerism (>1% engraftment), trafficked maternal immune cells, and immune responsiveness to donor cells. A total of 41.0% of maternal IUT recipients (mIUT) were chimeras (mean donor chimerism 3.0 ± 1.3%) versus 75.0% of paternal IUT recipients (pIUT, 3.6 ± 1.1%). Chimeras showed higher maternal microchimerism of CD4, CD8, and CD19 than non-chimeras. These maternal cells showed minimal responsiveness to B6 or BALB/c stimulation. To interrogate tolerance, mIUT were injected postnatally with 5E+6 B6 cells/pup; pIUT received BALB/c cells. IUT-treated pups showed no changes in trafficked maternal or fetal immune cell levels compared to controls. Donor-specific IgM and IgG were expressed by 1%-3% of recipients. mIUT splenocytes showed greater proliferation of regulatory T cells (Treg) upon BALB/c stimulation, while B6 stimulation upregulated the pro-inflammatory cytokines more than BALB/c. pIUT splenocytes produced identical Treg and cytokine responses to BALB/c and B6 cells, with higher Treg activity and lower pro-inflammatory cytokine expression upon exposure to BALB/c. In contrast, naïve fetal splenocytes demonstrated greater alloresponsiveness to BALB/c compared to B6 cells. Thus pIUT, associated with increased maternal cell trafficking, modulates fetal Treg, and cytokine responsiveness to donor cells more efficiently than mIUT, resulting in improved engraftment. Paternal donor cells may be considered alternatively to maternal donor cells for intrauterine and postnatal transplantation to induce tolerance and maintain engraftment.


Assuntos
Transplante de Medula Óssea , Sobrevivência de Enxerto/imunologia , Tolerância Imunológica/imunologia , Transplante Homólogo , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/imunologia , Transplante de Medula Óssea/métodos , Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas/métodos , Camundongos , Camundongos Endogâmicos BALB C , Quimeras de Transplante/imunologia , Transplante Homólogo/métodos
17.
Methods Mol Biol ; 2286: 95-105, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33534112

RESUMO

Bone marrow stromal cells (BMSCs, also known as bone marrow mesenchymal stem cells) are a plastic-adherent heterogeneous cell population that contain inherent skeletal progenitors and a subset of multipotential skeletal stem cells (SSCs). Application of BMSCs in therapeutic protocols implies its isolation and expansion under good manufacturing practices (GMP). Here we describe the procedures we have found to successfully generate practical BMSCs numbers, with preserved biological potency.


Assuntos
Tecnologia Biomédica/normas , Células da Medula Óssea/citologia , Osso e Ossos/citologia , Cultura Primária de Células/métodos , Antígenos CD34/genética , Antígenos CD34/metabolismo , Tecnologia Biomédica/métodos , Células Cultivadas , Técnicas de Cocultura/economia , Técnicas de Cocultura/métodos , Técnicas de Cocultura/normas , Custos e Análise de Custo , Meios de Cultura Livres de Soro/química , Humanos , Guias de Prática Clínica como Assunto , Cultura Primária de Células/economia , Cultura Primária de Células/normas , Células Estromais/citologia , Células Estromais/metabolismo
18.
Proc Natl Acad Sci U S A ; 118(6)2021 02 09.
Artigo em Inglês | MEDLINE | ID: mdl-33547233

RESUMO

Intracellular delivery of messenger RNA (mRNA)-based cancer vaccine has shown great potential to elicit antitumor immunity. To achieve robust antitumor efficacy, mRNA encoding tumor antigens needs to be efficiently delivered and translated in dendritic cells with concurrent innate immune stimulation to promote antigen presentation. Here, by screening a group of cationic lipid-like materials, we developed a minimalist nanovaccine with C1 lipid nanoparticle (LNP) that could efficiently deliver mRNA in antigen presenting cells with simultaneous Toll-like receptor 4 (TLR4) activation and induced robust T cell activation. The C1 nanovaccine entered cells via phagocytosis and showed efficient mRNA-encoded antigen expression and presentation. Furthermore, the C1 lipid nanoparticle itself induced the expression of inflammatory cytokines such as IL-12 via stimulating TLR4 signal pathway in dendritic cells. Importantly, the C1 mRNA nanovaccine exhibited significant antitumor efficacy in both tumor prevention and therapeutic vaccine settings. Overall, our work presents a C1 LNP-based mRNA cancer nanovaccine with efficient antigen expression as well as self-adjuvant property, which may provide a platform for developing cancer immunotherapy for a wide range of tumor types.


Assuntos
Antineoplásicos/imunologia , Lipídeos/química , RNA Mensageiro/administração & dosagem , RNA Mensageiro/imunologia , Transdução de Sinais , Receptor 4 Toll-Like/metabolismo , Animais , Células da Medula Óssea/citologia , Citocinas/metabolismo , Células Dendríticas/imunologia , Endocitose , Feminino , Células HEK293 , Humanos , Imunidade Inata , Ativação Linfocitária/imunologia , Melanoma Experimental/imunologia , Melanoma Experimental/patologia , Camundongos Endogâmicos C57BL , Nanopartículas/química , Linfócitos T/imunologia , Distribuição Tecidual
19.
Stem Cells Transl Med ; 10(6): 883-894, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33527780

RESUMO

While mesenchymal stromal cells are an appealing therapeutic option for a range of clinical applications, their potential to induce clotting when used systemically remains a safety concern, particularly in hypercoagulable conditions, such as in patients with severe COVID-19, trauma, or cancers. Here, we tested a novel preclinical approach aimed at improving the safety of mesenchymal stromal cell (MSC) systemic administration by use of a bioreactor. In this system, MSCs are seeded on the exterior of a hollow-fiber filter, sequestering them behind a hemocompatible semipermeable membrane with defined pore-size and permeability to allow for a molecularly defined cross talk between the therapeutic cells and the whole blood environment, including blood cells and signaling molecules. The potential for these bioreactor MSCs to induce clots in coagulable plasma was compared against directly injected "free" MSCs, a model of systemic administration. Our results showed that restricting MSCs exposure to plasma via a bioreactor extends the time necessary for clot formation to occur when compared with "free" MSCs. Measurement of cell surface data indicates the presence of known clot inducing factors, namely tissue factor and phosphatidylserine. Results also showed that recovering cells and flushing the bioreactor prior to use further prolonged clot formation time. Furthermore, application of this technology in two in vivo models did not require additional heparin in fully anticoagulated experimental animals to maintain target activated clotting time levels relative to heparin anticoagulated controls. Taken together the clinical use of bioreactor housed MSCs could offer a novel method to control systemic MSC exposure and prolong clot formation time.


Assuntos
Reatores Biológicos , COVID-19/terapia , Técnicas de Cultura de Células/métodos , Transplante de Células-Tronco Mesenquimais/métodos , Trombose/prevenção & controle , Animais , Anticoagulantes/farmacologia , Testes de Coagulação Sanguínea , Células da Medula Óssea/citologia , Células Cultivadas , Cães , Heparina/farmacologia , Humanos , Masculino , Membranas Artificiais , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/metabolismo , Infarto do Miocárdio/patologia , Infarto do Miocárdio/prevenção & controle , SARS-CoV-2 , Suínos
20.
Biomed Res Int ; 2021: 6697810, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33628811

RESUMO

Objective: To identify the shared genetic and epigenetic mechanisms between the osteogenic differentiation of dental pulp stem cells (DPSC) and bone marrow stem cells (BMSC). Materials and Methods: The profiling datasets of miRNA expression in the osteogenic differentiation of mesenchymal stem cells from the dental pulp (DPSC) and bone marrow (BMSC) were searched in the Gene Expression Omnibus (GEO) database. The differential expression analysis was performed to identify differentially expressed miRNAs (DEmiRNAs) dysregulated in DPSC and BMSC osteodifferentiation. The target genes of the DEmiRNAs that were dysregulated in DPSC and BMSC osteodifferentiation were identified, followed by the identification of the signaling pathways and biological processes (BPs) of these target genes. Accordingly, the DEmiRNA-transcription factor (TFs) network and the DEmiRNAs-small molecular drug network involved in the DPSC and BMSC osteodifferentiation were constructed. Results: 16 dysregulated DEmiRNAs were found to be overlapped in the DPSC and BMSC osteodifferentiation, including 8 DEmiRNAs with a common expression pattern (8 upregulated DEmiRNAs (miR-101-3p, miR-143-3p, miR-145-3p/5p, miR-19a-3p, miR-34c-5p, miR-3607-3p, miR-378e, miR-671-3p, and miR-671-5p) and 1 downregulated DEmiRNA (miR-671-3p/5p)), as well as 8 DEmiRNAs with a different expression pattern (i.e., miR-1273g-3p, miR-146a-5p, miR-146b-5p, miR-337-3p, miR-382-3p, miR-4508, miR-4516, and miR-6087). Several signaling pathways (TNF, mTOR, Hippo, neutrophin, and pathways regulating pluripotency of stem cells), transcription factors (RUNX1, FOXA1, HIF1A, and MYC), and small molecule drugs (curcumin, docosahexaenoic acid (DHA), vitamin D3, arsenic trioxide, 5-fluorouracil (5-FU), and naringin) were identified as common regulators of both the DPSC and BMSC osteodifferentiation. Conclusion: Common genetic and epigenetic mechanisms are involved in the osteodifferentiation of DPSCs and BMSCs.


Assuntos
Células da Medula Óssea/metabolismo , Diferenciação Celular , Bases de Dados de Ácidos Nucleicos , Polpa Dentária/metabolismo , Epigênese Genética , Osteogênese , Células-Tronco/metabolismo , Células da Medula Óssea/citologia , Polpa Dentária/citologia , Humanos , Células-Tronco/citologia
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