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1.
Gen Physiol Biophys ; 40(6): 551-559, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34897026

RESUMO

Neurodegenerative diseases are common problem for companion animals. Due to the limited ability of injured axons to regenerate, innovative therapies combined with rehabilitation have been applied and evaluated. Among them, stem cells and their conditioned media implantation, which can ameliorate damaged tissue has been suggested as a promising treatment strategy. The main goal of our study was to characterize mesenchymal stem cells (MSC) derived from canine adipose tissue (AT-MSC) and umbilical cord (UC-MSC) and analyse effect of their conditioned media (CM) on neurite outgrowth of neural progenitor cells isolated from the brain cortex of neonatal rats. MSC from both sources showed high osteogenic and chondrogenic potential and expression of CD90 and CD29. Furthermore, both UC-MSCCM and AT-MSCCM stimulated neurite growth. Interestingly, this effect was more pronounced with UC-MSCCM when compared to AT-MSCCM in vitro, which may be related to the different content of neurotrophic factors included in the CM.


Assuntos
Células-Tronco Mesenquimais , Células-Tronco Neurais , Animais , Diferenciação Celular , Meios de Cultivo Condicionados , Cães , Ratos , Cordão Umbilical
2.
Gen Physiol Biophys ; 40(6): 561-568, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34897027

RESUMO

Diffusion tensor imaging (DTI) is a magnetic resonance imaging technique used to characterize fibrous structures such as white matter in the central nervous system, including normal and spinal cord injury (SCI) conditions. Our aim was to evaluate the effect of alginate treatment in the rat SCI by DTI parametric measures. Ex vivo DTI data were collected by spin echo sequence with following parameters TR/TE: 2500 ms/32 ms and b-value of 1500 s/mm2. Main significant changes were found in fractional anisotropy (FA), and radial diffusivity (RD), between the saline- and alginatetreated group at the level of individual sections and whole spinal cord. Results indicate that ex vivo DTI can be used as a tool for tissue structure characterisation and both FA and RD as promising prognostic parameters of SCI treatment.


Assuntos
Imagem de Tensor de Difusão , Traumatismos da Medula Espinal , Alginatos , Animais , Imageamento por Ressonância Magnética , Ratos , Traumatismos da Medula Espinal/diagnóstico por imagem , Traumatismos da Medula Espinal/tratamento farmacológico
3.
Inflammation ; 44(6): 2419-2428, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34327573

RESUMO

Lactobacillus casei (L. casei) is one of the probiotic strains that may influence intestinal injury and inflammation in nonspecific intestinal diseases. We aimed to evaluate the effect of cell-free Lactobacillus casei 21L10 supernatant (LC) on the cell line HT-29 challenged with lipopolysaccharide (LPS) in order to modulate production of NO, cell proliferation, and apoptosis. Cell line HT-29 was stimulated with LPS in the presence or absence of LC. Our results showed that LC from L. casei 21L10 did not affect the viability of unstimulated HT-29 cells line. HT-29 cell line treatment with LC caused significant decrease of LPS induced NO production after 3 h, and 24 h, but not after 48 h. Proliferation activity of LPS stimulated HT-29 cell line analysed with MTT assay significantly decreased after 24 h and 48 h, but not after 3 h. The majority of LPS stimulated HT-29 cell line treated with LC showed annexin V/PI positivity at 48 h survival, which corresponded to late apoptotic/necrotic cell features. The observed differences suggest that cell-free L. casei 21L10 supernatant could participate in attenuation of LPS-induced inflammation, and may exhibit anti-proliferative and pro-apoptotic/necrotic effects. This study provides pilot data for the further development of L. casei exoproducts as an anti-inflammatory or anti-proliferative agent for the treatment of inflammatory and cancer diseases in gut. However, more data is needed before final conclusions of L. casei cell-free supernatant's efficacy can be drawn.


Assuntos
Anti-Inflamatórios/farmacologia , Antineoplásicos/farmacologia , Colo/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Doenças Inflamatórias Intestinais/tratamento farmacológico , Lacticaseibacillus casei/metabolismo , Anti-Inflamatórios/metabolismo , Antineoplásicos/metabolismo , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Colo/metabolismo , Colo/patologia , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , Células HT29 , Humanos , Mediadores da Inflamação/metabolismo , Doenças Inflamatórias Intestinais/metabolismo , Doenças Inflamatórias Intestinais/patologia , Lipopolissacarídeos/toxicidade , Óxido Nítrico/metabolismo , Fatores de Tempo
4.
Microorganisms ; 8(10)2020 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-32993120

RESUMO

The present study investigated the in vitro antibacterial, antibiofilm and anti-Quorum Sensing (anti-QS) activities of canine bone marrow mesenchymal stem cell-conditioned media (cBM MSC CM) containing all secreted factors <30 K, using a disc diffusion test (DDT), spectrophotometric Crystal Violet Assay (SCVA) and Bioluminescence Assay (BA) with QS-reporter Escherichia coli JM109 pSB1142. The results show a sample-specific bacterial growth inhibition (zones varied between 7-30 mm), statistically significant modulation of biofilm-associated Staphylococcus aureus and Escherichia coli bioluminescence (0.391 ± 0.062 in the positive control to the lowest 0.150 ± 0.096 in the experimental group, cf. 11,714 ± 1362 to 7753 ± 700, given as average values of absorbance A550 ± SD versus average values of relative light units to growth RLU/A550 ± SD). The proteomic analysis performed in our previous experiment revealed the presence of several substances with documented antibacterial, antibiofilm and immunomodulatory properties (namely, apolipoprotein B and D; amyloid-ß peptide; cathepsin B; protein S100-A4, galectin 3, CLEC3A, granulin, transferrin). This study highlights that cBM MSC CM may represent an important new approach to managing biofilm-associated and QS signal molecule-dependent bacterial infections. To the best of our knowledge, there is no previous documentation of canine BM MSC CM associated with in vitro antibiofilm and anti-QS activity.

5.
J Extracell Vesicles ; 9(1): 1727637, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32158520

RESUMO

Combining proteomics and systems biology approaches, we demonstrate that neonatal microglial cells derived from two different CNS locations, cortex and spinal cord, and cultured in vitro displayed different phenotypes upon different physiological or pathological conditions. These cells also exhibited greater variability in terms of cellular and small extracellular vesicles (sEVs) protein content and levels. Bioinformatic data analysis showed that cortical microglia exerted anti-inflammatory and neurogenesis/tumorigenesis properties, while the spinal cord microglia were more inflammatory. Interestingly, while both sEVs microglia sources enhanced growth of DRGs processes, only the spinal cord-derived sEVs microglia under LPS stimulation significantly attenuated glioma proliferation. These results were confirmed using the neurite outgrowth assay on DRGs cells and glioma proliferation analysis in 3D spheroid cultures. Results from these in vitro assays suggest that the microglia localized at different CNS regions can ensure different biological functions. Together, this study indicates that neonatal microglia locations regulate their physiological and pathological functional fates and could affect the high prevalence of brain vs spinal cord gliomas in adults.

6.
Neurochem Res ; 45(1): 134-143, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31006093

RESUMO

Spinal cord injury (SCI) often leads to irreversible neuro-degenerative changes with life-long consequences. While there is still no effective therapy available, the results of past research have led to improved quality of life for patients suffering from partial or permanent paralysis. In this review we focus on the need, importance and the scientific value of experimental animal models simulating SCI in humans. Furthermore, we highlight modern imaging tools determining the location and extent of spinal cord damage and their contribution to early diagnosis and selection of appropriate treatment. Finally, we focus on available cellular and acellular therapies and novel combinatory approaches with exosomes and active biomaterials. Here we discuss the efficacy and limitations of adult mesenchymal stem cells which can be derived from bone marrow, adipose tissue or umbilical cord blood and its Wharton's jelly. Special attention is paid to stem cell-derived exosomes and smart biomaterials due to their special properties as a delivery system for proteins, bioactive molecules or even genetic material.


Assuntos
Modelos Animais de Doenças , Neuroimagem/métodos , Traumatismos da Medula Espinal/diagnóstico por imagem , Traumatismos da Medula Espinal/terapia , Animais , Humanos , Imageamento por Ressonância Magnética/métodos , Procedimentos Neurocirúrgicos/métodos , Tomografia por Emissão de Pósitrons/métodos , Traumatismos da Medula Espinal/patologia , Transplante de Células-Tronco/métodos , Tomografia Computadorizada por Raios X/métodos , Resultado do Tratamento
8.
Int J Mol Sci ; 19(3)2018 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-29543759

RESUMO

It was recently shown that the conditioned medium (CM) of mesenchymal stem cells can enhance viability of neural and glial cell populations. In the present study, we have investigated a cell-free approach via CM from rat bone marrow stromal cells (MScCM) applied intrathecally (IT) for spinal cord injury (SCI) recovery in adult rats. Functional in vitro test on dorsal root ganglion (DRG) primary cultures confirmed biological properties of collected MScCM for production of neurosphere-like structures and axon outgrowth. Afterwards, rats underwent SCI and were treated with IT delivery of MScCM or vehicle at postsurgical Days 1, 5, 9, and 13, and left to survive 10 weeks. Rats that received MScCM showed significantly higher motor function recovery, increase in spared spinal cord tissue, enhanced GAP-43 expression and attenuated inflammation in comparison with vehicle-treated rats. Spared tissue around the lesion site was infiltrated with GAP-43-labeled axons at four weeks that gradually decreased at 10 weeks. Finally, a cytokine array performed on spinal cord extracts after MScCM treatment revealed decreased levels of IL-2, IL-6 and TNFα when compared to vehicle group. In conclusion, our results suggest that molecular cocktail found in MScCM is favorable for final neuroregeneration after SCI.


Assuntos
Meios de Cultivo Condicionados/farmacologia , Células-Tronco Mesenquimais/metabolismo , Regeneração Nervosa , Traumatismos da Medula Espinal/terapia , Animais , Células Cultivadas , Citocinas/genética , Citocinas/metabolismo , Gânglios Espinais/citologia , Masculino , Crescimento Neuronal/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Ratos , Ratos Wistar
9.
Spine J ; 13(12): 1881-91, 2013 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24012427

RESUMO

BACKGROUND CONTEXT: In recent years, hypothermia has been described as a therapeutic approach that leads to potential protective effects via minimization of secondary damage consequences, reduction of neurologic deficit, and increase of motor performance after spinal cord injury (SCI) in animal models and humans. PURPOSE: The objective of this study was to determine the therapeutic efficacy of hypothermia treatment on sensory-motor function and bladder activity outcome correlated with the white and gray matter sparing and neuronal survival after SCI in adult rats. STUDY DESIGN: A standardized animal model of compression SCI was used to test the hypothesis that hypothermia could have a neuroprotective effect on neural cell death and loss of white and/or gray matter. METHODS: Animals underwent spinal cord compression injury at the Th8-Th9 level followed by systemic hypothermia of 32.0°C with gradual re-warming to 37.0°C. Motor function of hind limbs (BBB score) and mechanical allodynia (von Frey hair filaments) together with function of urinary bladder was monitored in all experimental animals throughout the whole survival period. RESULTS: Present results showed that hypothermia had beneficial effects on urinary bladder activity and on locomotor function recovery at Days 7 and 14 post-injury. Furthermore, significant increase of NeuN-positive neuron survival within dorsal and ventral horns at Days 7, 14, and 21 were documented. CONCLUSIONS: Our conclusions suggest that hypothermia treatment may not only promote survival of neurons, which can have a significant impact on the improvement of motor and vegetative functions, but also induce mechanical allodynia.


Assuntos
Hiperalgesia/fisiopatologia , Hipotermia Induzida , Atividade Motora/fisiologia , Recuperação de Função Fisiológica/fisiologia , Traumatismos da Medula Espinal/fisiopatologia , Animais , Modelos Animais de Doenças , Imuno-Histoquímica , Masculino , Ratos , Ratos Wistar
10.
Cell Mol Neurobiol ; 29(6-7): 999-1013, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19350385

RESUMO

Ependymal cells (EC) in the spinal cord central canal (CC) are believed to be responsible for the postnatal neurogenesis following pathological or stimulatory conditions. In this study, we have analyzed the proliferation of the CC ependymal progenitors in adult rats processed to compression SCI or enhanced physical activity. To label dividing cells, a single daily injection of Bromo-deoxyuridine (BrdU) was administered over a 14-day-survival period. Systematic quantification of BrdU-positive ependymal progenitors was performed by using stereological principles of systematic, random sampling, and optical Dissector software. The number of proliferating BrdU-labeled EC increased gradually with the time of survival after both paradigms, spinal cord injury, or increased physical activity. In the spinal cord injury group, we have found 4.9-fold (4 days), 7.1-fold (7 days), 4.9-fold (10 days), and 5.6-fold (14 days) increase of proliferating EC in the rostro-caudal regions, 4 mm away from the epicenter. In the second group subjected to enhanced physical activity by running wheel, we have observed 2.1-2.6 fold increase of dividing EC in the thoracic spinal cord segments at 4 and 7 days, but no significant progression at 10-14 days. Nestin was rapidly induced in the ependymal cells of the CC by 2-4 days and expression decreased by 7-14 days post-injury. Double immunohistochemistry showed that dividing cells adjacent to CC expressed astrocytic (GFAP, S100beta) or nestin markers at 14 days. These data demonstrate that SCI or enhanced physical activity in adult rats induces an endogenous ependymal cell response leading to increased proliferation and differentiation primarily into macroglia or cells with nestin phenotype.


Assuntos
Células-Tronco Adultas/fisiologia , Epêndima/fisiologia , Epêndima/fisiopatologia , Compressão da Medula Espinal/fisiopatologia , Traumatismos da Medula Espinal/fisiopatologia , Animais , Bromodesoxiuridina , Contagem de Células , Proliferação de Células , Imuno-Histoquímica , Masculino , Atividade Motora , Ratos , Ratos Wistar , Canal Medular/fisiologia , Canal Medular/fisiopatologia , Vértebras Torácicas
11.
Cell Mol Neurobiol ; 26(7-8): 1167-80, 2006.
Artigo em Inglês | MEDLINE | ID: mdl-16897366

RESUMO

Human mesenchymal stem cells (hMSCs) derived from adult bone marrow represent a potentially useful source of cells for cell replacement therapy after nervous tissue damage. They can be expanded in culture and reintroduced into patients as autografts or allografts with unique immunologic properties. The aim of the present study was to investigate (i) survival, migration, differentiation properties of hMSCs transplanted into non-immunosuppressed rats after spinal cord injury (SCI) and (ii) impact of hMSC transplantation on functional recovery. Seven days after SCI, rats received i.v. injection of hMSCs (2x10(6) in 0.5 mL DMEM) isolated from adult healthy donors. Functional recovery was assessed by Basso-Beattie-Bresnahan (BBB) score weekly for 28 days. Our results showed gradual improvement of locomotor function in transplanted rats with statistically significant differences at 21 and 28 days. Immunocytochemical analysis using human nuclei (NUMA) and BrdU antibodies confirmed survival and migration of hMSCs into the injury site. Transplanted cells were found to infiltrate mainly into the ventrolateral white matter tracts, spreading also to adjacent segments located rostro-caudaly to the injury epicenter. In double-stained preparations, hMSCs were found to differentiate into oligodendrocytes (APC), but not into cells expressing neuronal markers (NeuN). Accumulation of GAP-43 regrowing axons within damaged white matter tracts after transplantation was observed. Our findings indicate that hMSCs may facilitate recovery from spinal cord injury by remyelinating spared white matter tracts and/or by enhancing axonal growth. In addition, low immunogenicity of hMSCs was confirmed by survival of donor cells without immunosuppressive treatment.


Assuntos
Transplante de Células-Tronco Mesenquimais , Traumatismos da Medula Espinal/terapia , Medula Espinal/fisiologia , Adulto , Animais , Comportamento Animal , Diferenciação Celular , Células Cultivadas , Feminino , Sobrevivência de Enxerto , Humanos , Imuno-Histoquímica , Masculino , Células-Tronco Mesenquimais/citologia , Pessoa de Meia-Idade , Regeneração Nervosa , Ratos , Ratos Wistar , Traumatismos da Medula Espinal/reabilitação , Transplante Heterólogo
12.
Neurochem Res ; 31(8): 1011-20, 2006 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16865557

RESUMO

The phenomenon of neuronal transdifferentiation performed on bone marrow mesenchymal stem cells (MSCs) has been criticized by recent studies indicating that acquired neuron-like morphology of induced MSCs is caused by cellular stress. Therefore, to test this hypothesis we have investigated whether exposure of rat MSCs (rMSCs) to chemical inducer 2 mM beta-mercaptoethanol (BME) for 1-3 h followed by 24 h incubation leads to HSP72 synthesis, thus suggesting higher resistance of rMSCs to oxidative damage. Present data from immunohistochemistry clearly indicate development of time-dependent sub-cellular HSP72 distribution, initially seen in nuclei at 1 h followed by its translocation to surrounding central cytoplasm and processes at 2-3 h after BME stimulation. Western blot (WB) analysis confirmed the expression of HSP72 protein in induced rMSCs at both stimulation periods. Furthermore, preconditioned rMSCs with BME for 1 h expressing HSP72 positivity at 24 h showed higher resistance (78 +/- 10% of survival cells) to oxidative stress caused by 1 mM H(2)O(2) when compared to those preconditioned for 3 h (59 +/- 8% of survival cells) or control-unconditioned rMSCs exposed to the same stressor conditions (56 +/- 6% of survival cells). Thus, the cellular protection was lost if the duration of BME preconditioning was increased as far as possible (3 h) (while still remaining sub-lethal). This suggests that exposure of rMSCs to the optimal concentration of BME (2 mM) during optimal induction period (1 h) mediate their protection and increases resistance to oxidative injury, while over crossing these limits is in-effective. In addition, our findings confirm that cultured rMSCs remain competent to be preconditioned by BME, through a pathway that may increase the antioxidant balance or involve activation of HSP72 protein induced tolerance.


Assuntos
Diferenciação Celular/fisiologia , Proteínas de Choque Térmico HSP72/metabolismo , Células-Tronco Mesenquimais/citologia , Células-Tronco Mesenquimais/fisiologia , Estresse Oxidativo , Regulação para Cima , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/efeitos dos fármacos , Células da Medula Óssea/fisiologia , Forma Celular , Sobrevivência Celular , Células Cultivadas , Peróxido de Hidrogênio/farmacologia , Masculino , Mercaptoetanol/farmacologia , Células-Tronco Mesenquimais/efeitos dos fármacos , Oxidantes/farmacologia , Ratos , Ratos Wistar
13.
Berl Munch Tierarztl Wochenschr ; 117(3-4): 145-7, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15046462

RESUMO

Two strains of lactobacilli (Lactobacillus acidophilus T-135 and Lactobacillus plantarum 4/97) were selected in order to study their inhibitory properties against frequent udder pathogens (Escherichia coli, Staphylococcus aureus, Streptococcus agalactiae, Streptococcus uberis, Salmonella enteritidis and Bacillus pumilus), their production of organic acids as well as their ability to survive on the teat skin, the teat duct mucosa and in a lipoid emulsion. Both strains inhibited the tested pathogenic microbes and survived on the investigated surfaces and in an emulsion for more than 6 hours and 11 days, respectively.


Assuntos
Antibiose , Lactobacillus acidophilus/fisiologia , Lactobacillus/fisiologia , Glândulas Mamárias Animais/microbiologia , Probióticos , Animais , Bacillus/crescimento & desenvolvimento , Bovinos , Indústria de Laticínios/métodos , Escherichia coli/crescimento & desenvolvimento , Feminino , Salmonella enteritidis/crescimento & desenvolvimento , Staphylococcus aureus/crescimento & desenvolvimento , Streptococcus/crescimento & desenvolvimento
14.
Acta Histochem ; 104(4): 381-5, 2002.
Artigo em Inglês | MEDLINE | ID: mdl-12553707

RESUMO

We have characterized segmental and laminar distribution patterns of Fos-immunopositive (Fos-IP) neurons in spinal cord segments L3-L6 after carrageenan treatment. A large number of Fos-IP neurons was found in the medial region of the ipsilateral dorsal horn laminae I-II at 4 and 6 h postinjection (pi). At one day pi, the number of Fos-IP neurons was decreased significantly, which correlated with suppression of inflammation in the affected hind paw. Bilaterally, Fos-IP neurons reappeared in the L3-L6 spinal cord segments at 3-4 days pi, mainly in the deep laminae LV-LVI. However, signs of inflammation had distinctly attenuated. Our data indicate a biphasic trend in Fos-IP in this experimental model of inflammation.


Assuntos
Carragenina , Proteínas Proto-Oncogênicas c-fos/metabolismo , Medula Espinal/metabolismo , Animais , Carragenina/farmacologia , Membro Posterior/efeitos dos fármacos , Membro Posterior/patologia , Inflamação/induzido quimicamente , Inflamação/metabolismo , Inflamação/patologia , Masculino , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Neurônios/patologia , Ratos , Ratos Wistar , Medula Espinal/efeitos dos fármacos , Medula Espinal/patologia
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