RESUMO
Nocardia seriolae is a severe bacterial pathogen that has seriously affected the development of aquaculture industry. Largemouth bass (Micropterus salmoides) is a commercially significant freshwater fish that suffers a variety of environmental threats, including bacterial pathogens. However, the immune responses and metabolic alterations of largemouth bass to N. seriolae infection remain largely unclear. We discovered that N. seriolae caused pathological alterations in largemouth bass and shifted the transcript of immune-related and apoptotic genes in head kidney after infection. To answer the aforementioned question, a combined transcriptome and metabolome analysis was employed to explore the alterations in genes, metabolites, and metabolic pathways in largemouth bass following bacterial infection. A total of 3579 genes and 1929 metabolites are significant differentially changed in the head kidney post infection. In response to N. seriolae infection, host modifies the PI3K-Akt signaling pathway, TCA cycle, glycolysis, and amino acid metabolism. The integrated analysis of transcriptome and metabolome suggested that with the arginine metabolism pathway as the core, multiple biomarkers (arg gene, arginine) are involved in the antibacterial and immune functions of largemouth bass. Thus, we hypothesized that arginine plays a crucial role in the immune responses of largemouth bass against N. seriolae infection, and increasing arginine levels suitably is beneficial for the host against bacterial infection. Our results shed light on the regulatory mechanism of largemouth bass resistance to N. seriolae infection and contributed to the development of more effective N. seriolae resistance strategies.
Assuntos
Infecções Bacterianas , Bass , Nocardiose , Nocardia , Animais , Transcriptoma , Fosfatidilinositol 3-Quinases/genética , Metaboloma , ArgininaRESUMO
Investigation on the physiochemical nature involved in the production of fatty acid catalyzed by the vesicles is of importance to understand the digestion of lipid. In this paper, the effects of crowding degree, which was constructed by polyethylene glycol (PEG), on the autocatalytic production of fatty acid with different chain lengths was studied. The results showed that the higher crowding degree led to the slower production rate of decanoic acid but the faster rate of oleic acid. The reason lies in that the presence of macromolecules resulted in the increased sizes of decanoic acid vesicles, but decreased sizes of oleic acid vesicles. Meanwhile, decanoic acid vesicles in more crowded medium exhibited viscous behavior, whereas oleic acid displayed elastic behavior. This research provides useful information for understanding the unusual autocatalyzed production of fatty acid in macromolecular crowding and may also draw an attention to the physiologically relevant lipid digestion.
RESUMO
The erroneous differentiation of tendon-derived stem cells (TDSCs) into adipocytes, chondrocytes, and osteoblasts is believed to play an important role in the development of tendinopathy. However, the regulatory mechanisms of TDSC differentiation remain unclear. The aim of this study is to investigate the contribution and mechanism of the tendon microstructural disruption to the differentiation of TDSCs. Bovine Achilles tendons were sliced. The tendon slices were stretched with different tensile strains to mimic the tendon structure alteration at various scales. The TDSCs were cultured on the tendon slices. The differentiation of TDSCs and endoplasmic reticulum (ER) stress in the TDSCs were investigated with quantitative reverse transcription polymerase chain reaction, immunostaining and western blot. The effect of ER stress inhibition on chondrogenic differentiation of the TDSCs was further investigated. The structural alteration did not affect the viability of TDSCs. However, the structural alteration of tendon slices with 6.4% strain promoted TDSCs to express the chondrogenic genes. ER stress-related markers, ATF-4 and PERK, were also upregulated. With the inhibition of ER stress, the expression of ATF-4 and the chondrogenic gene SOX9 of TDSCs were inhibited. The study indicated that tendon microdamage could induce the chondrogenic differentiation of TDSCs through triggering ER stress to activate ATF-4 and SOX9 subsequently.
Assuntos
Tendinopatia , Tendões , Ratos , Animais , Bovinos , Ratos Sprague-Dawley , Diferenciação Celular , Células-Tronco , Tendinopatia/metabolismoRESUMO
The rat is frequently used as a model to study the characteristics, aetiology and pathology of the Achilles tendon. However, though the structure of the human Achilles tendon has been extensively investigated, the anatomical structure of the rat Achilles tendon remains unclear, which impedes the ability to use rats to study Achilles tendinopathy. The purpose of this study was to reveal the structure of the rat Achilles tendon and to explore its similarities with the human Achilles tendon through an anatomical dissection of 80 rat Achilles tendons (40 female, 40 male). This study found that the subtendons of the rat Achilles tendon originating from the triceps surae muscle were twisted, and each subtendon also had its own torsion. The extent of these two types of torsion could be very different between rats. Alterations in this torsion may result in distinct stress fields in the Achilles tendon, which may play a critical role in the pathogenesis of Achilles tendinopathy. This study provides an important basis to support the use of rats as model animals to investigate the characteristics of the human Achilles tendon and Achilles tendinopathy.
Assuntos
Tendão do Calcâneo , Tendinopatia , Animais , Dissecação , Feminino , Masculino , Músculo Esquelético , RatosRESUMO
Fibrin glue has been widely used as a surgical sealing and hemostatic agent. Its application is restricted due to poor tissue adhesion and low mechanical strength. To develop better tissue sealant and hemostatic agent, this study prepared the injectable hydrogels by chemically cross-linking gelatin (G) with or without hyaluronic acid (HA) in situ at a mild condition. The rheological analysis, Fourier transform infrared spectroscopy, swelling, proteolytic degradation, biocompatibility, tissue sealing, and hemostatic ability of the hydrogels were investigated. It was found that the chemical cross-linking rapidly formed in both self-crosslinking gelatin (sc-G) and gelatin/hyaluronate acid (G/HA) hydrogels. The hydrogels could be degraded by trypsin and had a desirable biocompatibility. The tissue sealing ability of the hydrogels was superior to fibrin glue. Furthermore, the G/HA hydrogel had similar hemostatic performance as fibrin glue, and was better than that of gelatin hydrogel. The results in the study indicated that the G/HA hydrogel could be used in clinic as a tissue sealant or surgical hemostat.
Assuntos
Materiais Biocompatíveis/química , Adesivo Tecidual de Fibrina/química , Gelatina/química , Ácido Hialurônico/química , Hidrogéis/química , Materiais Biocompatíveis/administração & dosagem , Materiais Biocompatíveis/efeitos adversos , Sobrevivência Celular , Reagentes de Ligações Cruzadas/química , Adesivo Tecidual de Fibrina/administração & dosagem , Adesivo Tecidual de Fibrina/efeitos adversos , Hemostasia , Hemostáticos/química , Hemostáticos/metabolismo , Humanos , Hidrogéis/administração & dosagem , Hidrogéis/efeitos adversos , Injeções , ReologiaRESUMO
Tissue sealants are used for hemorrhage control which is imperative in many surgical procedures. It is a highly challenging task to obtain the ideal tissue sealant. Only a few commercially tissue sealants are available to be used for internal tissue or organ hemorrhage control. This study introduced two in situ injectable hydrogels for hemorrhage control: self-crosslinking gelatin (sc-G) hydrogel and hyaluronic acid/gelatin (HA/G) hydrogel. They were prepared on the tissue surface in situ and characterized by rheological analysis, stability, cytotoxicity, and bursting strength test. The hemostatic ability of the hydrogels was evaluated in a liver-bleeding rat model. The sc-G and HA/G hydrogels gelled around 90â¯s and 50â¯s, respectively. They were preferable for cell attachment and proliferation. The bursting strengths of both hydrogels exceeded that of fibrin glue. The hemostatic ability of HA/G hydrogel was better than that of sc-G hydrogel, and was same as that of fibrin glue. The HA/G hydrogel could be used as a tissue sealant for hemorrhage control in clinic.
Assuntos
Ácido Hialurônico/química , Hidrogéis/química , Animais , Materiais Biocompatíveis/química , Gelatina/química , Hemostasia , Masculino , Ratos , Ratos Sprague-Dawley , Engenharia Tecidual/métodos , Alicerces Teciduais/químicaRESUMO
Tendinopathy is prevalent in athletic and many occupational populations; nevertheless, the pathogenesis of tendinopathy remains unclear. Tendon-derived stem cells (TDSCs) were regarded as the key culprit for the development of tendinopathy. However, it is uncertain how TDSCs differentiate into adipocytes, chondrocytes, or osteocytes in the degenerative microenvironment of tendinopathy. So in this study, the regulating effects of the degenerative tendon microenvironment on differentiation of TDSCs were investigated. TDSCs were isolated from rat Achilles tendons and were grown on normal and degenerative (prepared by stress-deprived culture) decellularized tendon slices (DTSs). Immunofluorescence staining, H&E staining, real-time PCR, and Western blot were used to delineate the morphology, proliferation, and differentiation of TDSCs in the degenerative microenvironment. It was found that TDSCs were much more spread on the degenerative DTSs than those on normal DTSs. The tenocyte-related markers, COL1 and TNMD, were highly expressed on normal DTSs than the degenerative DTSs. The expression of chondrogenic and osteogenic markers, COL2, SOX9, Runx2, and ALP, was higher on the degenerative DTSs compared with TDSCs on normal DTSs. Furthermore, phosphorylated FAK and ERK1/2 were reduced on degenerative DTSs. In conclusion, this study found that the degenerative tendon microenvironment induced TDSCs to differentiate into chondrogenic and osteogenic lineages. It could be attributed to the cell morphology changes and reduced FAK and ERK1/2 activation in the degenerative microenvironment of tendinopathy.
RESUMO
Tendon derived stem cells (TDSCs) were vital in tendon homeostasis. Nevertheless, the regulation of TDSCs differentiation in tendinopathy is unclear. Matrix stiffness modulated stem cells differentiation, and matrix stiffness of tendinopathic tissues decreased significantly. In order to clarify the role of matrix stiffness in TDSCs differentiation, they were cultured on the gelatin hydrogels with the stiffness from 2.34⯱â¯1.48â¯kPa to 24.09⯱â¯14.03â¯kPa. The effect of matrix stiffness on TDSCs proliferation and differentiation were investigated with CCK8 assay, immunofluorescences, real time PCR and western blot. It was found the proliferation of TDSCs increased and more stress fibers formed with increasing matrix stiffness. The differentiation of TDSCs into tenogenic, chondrogenic, and osteogenic lineages were inhibited on stiff hydrogel evidenced by reduced expression of tenocyte markers THBS4, TNMD, SCX, chondrocyte marker COL2, and osteocyte markers Runx2, Osterix, and ALP. Furthermore, the phosphorylation of FAK and ERK1/2 were enhanced when TDSCs grew on stiff hydrogel. After FAK or ERK1/2 was inhibited, the effect of matrix stiffness on differentiation of TDSCs was inhibited as well. The above results indicated matrix stiffness modulated the proliferation and differentiation of TDSCs, and the regulation effect could correlate to the activation of FAK or ERK1/2.
Assuntos
Diferenciação Celular , Quinase 1 de Adesão Focal/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Células-Tronco/enzimologia , Tendões/citologia , Citoesqueleto de Actina/ultraestrutura , Animais , Proliferação de Células , Sobrevivência Celular , Ativação Enzimática , Gelatina , Hidrogéis , Sistema de Sinalização das MAP Quinases , Ratos Sprague-Dawley , Células-Tronco/citologiaRESUMO
BACKGROUND: Postoperative intra-abdominal adhesion often causes many complications. Chitosan fluid has been used in clinic to prevent intra-abdominal adhesion. However, fluid can be easily diluted and cannot stay on the wound site. As hydrogel is able to form stable physical barrier to separate injured tissues, we developed a chitosan hydrogel for better prevention of intra-abdominal adhesion in this study. METHODS: We synthesized a carbodiimide-derivatized chitosan gelatin (cd-CS-gelatin) hydrogel and investigated its rheological properties. A rat model was used to compare the anti-adhesion effect of chitosan hydrogel and fluid. The wounds were created with damage of the underlying muscle of the abdominal wall and the serosal layer of the cecum. They were coated with chitosan fluid or cd-CS-gelatin hydrogel. At day 14 after surgery, the animals were euthanized and intra-abdominal adhesion was assessed. RESULTS: The cd-CS-gelatin hydrogel solidified within 3min after the mixing of the reagents. The cecum-abdomen adhesion occurred in all rats without anti-adhesion treatment. The application of cd-CS-gelatin significantly reduced the adhesion rate from 100% to 50%, compared the chitosan fluid only to 88%. The decrease of adhesion breaking strength also manifested that cd-CS-gelatin was more effective than chitosan fluid to reduce postsurgical intra-abdominal adhesion formation. CONCLUSIONS: Chitosan hydrogel is more effective than chitosan fluid to prevent postoperative cecum-abdomen adhesion. It indicates that hydrogel could be a more promising state than liquid to prevent postoperative intra-abdominal adhesion.
Assuntos
Hidrogéis/química , Adesivos , Animais , Carbodi-Imidas , Quitosana , Reagentes de Ligações Cruzadas , Ratos , Aderências TeciduaisRESUMO
BACKGROUND: Adhesions frequently occur after abdominal surgery. Many anti-adhesion products have been used in clinic. However, the evidences are short for surgeons to reasonably choose the suitable anti-adhesion produces in clinical practice. This study provided such evidence by comparing the efficiency of five products to prevent abdominal adhesion formation in a rat model. METHODS: Fifty-six Sprague-Dawley rats were randomly divided into seven groups: sham-operation group, adhesion group, and five product groups (n = 8). The abdomens of rats were opened. The injuries were created on abdominal wall and cecum in the adhesion and product groups. The wounds on abdominal wall and cecum of rats in the adhesion group were not treated before the abdomens were closed. The wounds on abdominal wall and cecum of rats in the product groups were covered with anti-adhesion product: polylactic acid (PLA) film, Seprafilm®, medical polyethylene glycol berberine liquid (PEG), medical sodium hyaluronate gel (HA), or medical chitosan (Chitosan). Fourteen days after surgery, the adhesions were evaluated by incidence, severity, adhesion area on abdominal wall and adhesion breaking strength. RESULTS: The application of PLA film and Seprafilm® significantly reduced the incidence, severity, adhesion area and breaking strength of cecum-abdomen adhesion (P<0.05). HA, PEG and Chitosan failed to significantly reduce the cecum-abdomen adhesion (P>0.05). The statistical significances in the incidence and severity of abdomen-adipose adhesion between adhesion group and the product groups were not achieved. However, Seprafilm® was more effective to reduce abdomen-adipose adhesion than PLA film. Furthermore, it was found that the products tested in this study did not effectively reduce cecum-adipose adhesion. The application of PEG could result in abdomen-small intestine adhesion. CONCLUSION: Based on the results of this study, the preference order of anti-adhesion products used to reduce postsurgical intra-abdominal adhesion formation is Seprafilm > PLA >> HA > Chitosan > PEG.
Assuntos
Parede Abdominal/cirurgia , Materiais Biocompatíveis/uso terapêutico , Quitosana/uso terapêutico , Ácido Hialurônico/uso terapêutico , Poliésteres/uso terapêutico , Complicações Pós-Operatórias/prevenção & controle , Aderências Teciduais/prevenção & controle , Animais , Ceco/cirurgia , Modelos Animais de Doenças , Feminino , Ratos , Ratos Sprague-Dawley , CicatrizaçãoRESUMO
BACKGROUND: Adhesion grades and adhesion breaking strength are widely used to assess severity of intraperitoneal adhesion in animal models. However, the results of adhesion grades have the large deviations due to vary personal experience. Adhesion breaking strength ignores the details of adhesion. This study introduced work of separation, the energy consumption during breakage of adhesion, to better evaluate intraperitoneal adhesion. METHODS: The intraperitoneal adhesion was induced by traumas created at rat cecum and adjacent abdominal wall. The wounds were coated with or without sodium hyaluronate. On day 14 after surgery, the intraperitoneal adhesion was assessed by adhesion density grade, adhesion area grade, adhesion breaking strength and work of separation. The healing of parietal peritoneum was evaluated with histology, adhesion breaking strength and work of separation. FINDINGS: The severity of adhesion evaluated with work of separation was consistent with those obtained from the grades of adhesion density, adhesion area and adhesion breaking strength. Work of separation had a linear correlation with adhesion breaking strength. Furthermore, the results of histological examination and work of separation demonstrated that adhesion significantly delayed healing process of abdominal wall muscles. INTERPRETATION: Work of separation can quantify all intraperitoneal adhesions rather than the major one by other methods. It is a more precise method to evaluate postoperative adhesions, especially those including adipose tissue. This study proved that work of separation could be a reliable method to assess intraperitoneal adhesion and tissue healing.
Assuntos
Doenças Peritoneais/fisiopatologia , Cicatrização/fisiologia , Parede Abdominal/fisiopatologia , Animais , Modelos Animais de Doenças , Feminino , Humanos , Ácido Hialurônico , Doenças Peritoneais/patologia , Ratos Sprague-Dawley , Aderências Teciduais/fisiopatologiaRESUMO
Bletilla striata has been used as traditional Chinese medicine for several centuries. In recent years, the quality and quantity of wild B. striata plants have declined sharply due to habitat deterioration and human over-exploitation. Therefore, it is of great urgency to evaluate and protect B. striata wild plant resource. In this study, sequence-related amplified polymorphism (SRAP) markers were applied to assess the level and pattern of genetic diversity in twelve populations of B. striata. The results showed a high level of genetic diversity (PPB = 90.48%, H = 0.349 4, I = 0.509 6) and moderate genetic differentiation among populations (G(st) = 0.260 9). Based on the unweighted pair-group method with arithmetic average (UPGMA), twelve populations gathered in three clusters. The cluster 1 included four populations. There are Nanjing, Zhenjiang, Xuancheng and Hangzhou. The seven populations which come from Hubei Province, Hunan Province, Jiangxi Province and Guizhou Province belonged to the cluster 2. The cluster 3 only contained Wenshan population. Moreover, Mantel test revealed significant positive correlation between genetic distances and geographic distances (r = 0.632 9; P < 0.000 1). According to the results, we proposed a series of conservation consideration for B. striata.
Assuntos
Variação Genética , Orchidaceae/genética , Filogenia , China , Marcadores Genéticos , Genética Populacional , Plantas Medicinais/genéticaRESUMO
BACKGROUND AND PURPOSE: Neuropeptide FF (NPFF) behaves as an endogenous opioid-modulating peptide. In the present study, the opioid and NPFF pharmacophore-containing chimeric peptide BN-9 was synthesized and pharmacologically characterized. EXPERIMENTAL APPROACH: Agonist activities of BN-9 at opioid and NPFF receptors were characterized in in vitro cAMP assays. Antinociceptive activities of BN-9 were evaluated in the mouse tail-flick and formalin tests. Furthermore, its side effects were investigated in rotarod, antinociceptive tolerance, reward and gastrointestinal transit tests. KEY RESULTS: BN-9 acted as a novel multifunctional agonist at µ, δ, κ, NPFF1 and NPFF2 receptors in cAMP assays. In the tail-flick test, BN-9 produced dose-related antinociception and was approximately equipotent to morphine; this antinociception was blocked by µ and κ receptor antagonists, but not by the δ receptor antagonist. In the formalin test, supraspinal administration of BN-9 produced significant analgesia. Notably, repeated administration of BN-9 produced analgesia without loss of potency over 8 days. In contrast, repeated i.c.v. co-administration of BN-9 with the NPFF receptor antagonist RF9 produced significant antinociceptive tolerance. Furthermore, i.c.v. BN-9 induced conditioned place preference. When given by the same routes, BN-9 had a more than eightfold higher ED50 value for gastrointestinal transit inhibition compared with the ED50 values for antinociception. CONCLUSIONS AND IMPLICATIONS: BN-9 produced a robust, nontolerance-forming analgesia with limited inhibition of gastrointestinal transit. As BN-9 is able to activate both opioid and NPFF systems, this provides an interesting approach for the development of novel analgesics with minimal side effects.
Assuntos
Analgésicos/farmacologia , Oligopeptídeos/farmacologia , Dor/tratamento farmacológico , Receptores de Neuropeptídeos/agonistas , Receptores Opioides/agonistas , Analgésicos/efeitos adversos , Analgésicos/química , Animais , Relação Dose-Resposta a Droga , Masculino , Camundongos , Camundongos Endogâmicos , Oligopeptídeos/efeitos adversos , Oligopeptídeos/química , Relação Estrutura-AtividadeRESUMO
Adhesions often occur after abdominal surgery. It could cause chronic pelvic pain, intestinal obstruction, and infertility. A hydrogel biomaterial, carbodiimide-derivatized hyaluronic acid gelatin (cd-HA gelatin), has been successfully used to reduce adhesion formation after flexor tendon grafting. This study investigated the efficacy of cd-HA gelatin in preventing postsurgical peritoneal adhesions in a rat model. The surgical traumas were created on the underlying muscle of the abdominal wall and the serosal layer of the cecum. The wounds were covered with or without cd-HA gelatin. Animals were euthanized at day 14 after surgery. Adhesion formation was assessed with adhesion degree and adhesion breaking strength. The healing of abdominal wall was evaluated with biomechanical testing and histological analysis. The adhesions occurred in all rats (n = 12) without cd-HA gelatin treatment. The application of cd-HA gelatin significantly reduced the adhesion rate from 100% to 58%. The decrease of adhesion breaking strength also manifested that cd-HA gelatin could reduce postsurgical intra-abdominal adhesion formation. Moreover, it was found that cd-HA gelatin was a safe material and could promote tissue healing. The cd-HA gelatin hydrogel could reduce the formation of intra-abdominal adhesions without adversely effects on wound healing.
Assuntos
Parede Abdominal/patologia , Parede Abdominal/cirurgia , Materiais Biocompatíveis/uso terapêutico , Gelatina/uso terapêutico , Ácido Hialurônico/uso terapêutico , Hidrogéis/uso terapêutico , Aderências Teciduais/prevenção & controle , Animais , Materiais Biocompatíveis/química , Carbodi-Imidas/química , Carbodi-Imidas/uso terapêutico , Feminino , Gelatina/química , Ácido Hialurônico/análogos & derivados , Hidrogéis/química , Ratos Sprague-Dawley , Tendões/transplante , Aderências Teciduais/etiologia , Aderências Teciduais/patologia , Cicatrização/efeitos dos fármacosRESUMO
In this study, 17 kinds of Dendrobium species of Fengdous including 39 individuals were collected from 4 provinces. Mitochondrial gene sequences co I, nad 5, nad 1-intron 2 and chloroplast gene sequences rbcL, matK amd psbA-trnH were amplified from these materials, as well as nrDNA ITS. Furthermore, suitable sequences for identification of Dendrobium species of Fengdous were screened by K-2-P and P-distance. The results showed that during the mentioned 7 sequences, nrDNA ITS, nad 1-intron 2 and psbA-trnH which had a high degree of variability could be used to identify Dendrobium species of Fengdous. However, single fragment could not be used to distinguish D. moniliforme and D. huoshanense. Moreover, compared to other combined fragments, new type combined fragments nrDNA ITS+nad 1-intron 2 was more effective in identifying the original plants of Dendrobium species and could be used to identify D. huoshanense and D. moniliforme. Besides, according to the UPGMA tree constructed with nrDNA ITS+nad 1-intron 2, 3 inspected Dendrobium plants were identified as D. huoshanense, D. moniliforme and D. officinale, respectively. This study identified Dendrobium species of Fengdous by combined fragments nrDNA ITS+nad 1-intron 2 for the first time, which provided a more effective basis for identification of Dendrobium species. And this study will be helpful for regulating the market of Fengdous.
Assuntos
DNA Espaçador Ribossômico/genética , Dendrobium/classificação , Genes de Cloroplastos , Íntrons , DNA de Plantas/genética , Dendrobium/genética , Genes de Plantas , Plantas Medicinais/classificação , Plantas Medicinais/genéticaRESUMO
Achilles tendon is one of the most commonly injured tendons. Mechanical force is regarded as a major causative factor. However, the biomechanics of Achilles tendon and mechanical mechanism of the injuries are unclear. Lubricin expresses at regions exposed to sliding motion and shear force in a number of tissues. This study investigated the distribution and concentration of lubricin in human Achilles tendons for better understanding the biomechanics of Achilles tendon. Achilles tendons were harvested from nine cadavers. Lubricin was extracted from various locations proximal to the calcaneal insertion and quantified with ELISA. The distribution of lubricin was investigated with immunohistochemistry. Lubricin was mainly identified at the interfaces of tendon fascicles, especially in the mid-portion of the tendon. The concentration of lubricin in Achilles tendons varied by individual and the distance from its calcaneal insertion. The distal portion of the tendon had a higher concentration of lubricin than the proximal regions of the tendon. This study suggests the presence of intratendinous sliding motion of fascicles and shear force at interfaces of fascicles in human Achilles tendon. Shear force could be an important mechanical factor for the development of Achilles tendinopathy and rupture.
Assuntos
Tendão do Calcâneo/fisiologia , Glicoproteínas/fisiologia , Tendão do Calcâneo/química , Adulto , Idoso , Feminino , Glicoproteínas/análise , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
Cell-based tissue engineered tendons have potential to improve clinical outcomes following rotator cuff repair, especially in large or massive rotator cuff tears, which pose a great clinical challenge. The aim of this study was to develop a method of constructing a functional engineered tendon patch for rotator cuff repair with cyclic mechanical stimulation. Decellularized tendon slices (DTSs) were seeded with BMSCs and subjected to cyclic stretching for 1, 3, or 7 days. The mechanical properties, morphologic characteristics and tendon-related gene expression of the constructs were investigated. Viable BMSCs were observed on the DTS after 7 days. BMSCs penetrated into the DTSs and formed dense cell sheets after 7 days of mechanical stretching. Gene expression of type I collagen, decorin, and tenomodulin significantly increased in cyclically stretched BMSC-DTS constructs compared with the unstrained control group (P < 0.05). The ultimate tensile strength and stiffness of the cyclically stretched tendon constructs were similar to the unstrained control group (P > 0.05). In conclusion, mechanical stimulation of BMSC-DTS constructs upregulated expression of tendon-related proteins, promoted cell tenogenic differentiation, facilitated cell infiltration and formation of cell sheets, and retained mechanical properties. The patch could be used as a graft to enhance the surgical repair of rotator cuff tears.
Assuntos
Células-Tronco Mesenquimais/citologia , Manguito Rotador/patologia , Estresse Mecânico , Engenharia Tecidual/métodos , Alicerces Teciduais/química , Cicatrização , Animais , Fenômenos Biomecânicos , Forma Celular , Sobrevivência Celular , Células Cultivadas , Cães , Proteínas da Matriz Extracelular/genética , Proteínas da Matriz Extracelular/metabolismo , Regulação da Expressão Gênica , Reação em Cadeia da Polimerase em Tempo Real , Manguito Rotador/ultraestruturaRESUMO
The diamagnetic levitation as a novel ground-based model for simulating a reduced gravity environment has recently been applied in life science research. In this study a specially designed superconducting magnet with a large gradient high magnetic field (LG-HMF), which can provide three apparent gravity levels (µ-g, 1-g, and 2-g), was used to simulate a space-like gravity environment. Osteocyte, as the most important mechanosensor in bone, takes a pivotal position in mediating the mechano-induced bone remodeling. In this study, the effects of LG-HMF on gene expression profiling of osteocyte-like cell line MLO-Y4 were investigated by Affymetrix DNA microarray. LG-HMF affected osteocyte gene expression profiling. Differentially expressed genes (DEGs) and data mining were further analyzed by using bioinfomatic tools, such as DAVID, iReport. 12 energy metabolism related genes (PFKL, AK4, ALDOC, COX7A1, STC1, ADM, CA9, CA12, P4HA1, APLN, GPR35 and GPR84) were further confirmed by real-time PCR. An integrated gene interaction network of 12 DEGs was constructed. Bio-data mining showed that genes involved in glucose metabolic process and apoptosis changed notablly. Our results demostrated that LG-HMF affected the expression of energy metabolism related genes in osteocyte. The identification of sensitive genes to special environments may provide some potential targets for preventing and treating bone loss or osteoporosis.
Assuntos
Metabolismo Energético , Osteócitos/fisiologia , Transcriptoma , Animais , Linhagem Celular , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Campos Magnéticos , Camundongos , Anotação de Sequência MolecularRESUMO
Hairpin DNA (hpDNA) as a novel biobarcode was conjugated with gold nanoparticles (AuNPs) and a reporter DNA (rpDNA) to form hpDNA/AuNP/rpDNA nanoparticles for the detection of an oligonucleotide sequence associated with Helicobacter pylori as a model target. The rpDNA is complementary to about a half-portion of the target DNA sequence (tDNA). A capture DNA probe (cpDNA), complementary to the other half of the tDNA, was immobilized on the surface of a gold electrode. In the presence of tDNA, a sandwich structure of (hpDNA/AuNP/rpDNA)/tDNA/cpDNA was formed on the electrode surface. The differential pulse voltammetry (DPV) detection was based on [Ru(NH3)5(3-(2-phenanthren-9-yl-vinyl)-pyridine)](2+), an electroactive complex that binds to the sandwich structure by its intercalation with the hpDNA and the double-stranded DNA (dsDNA) of the sandwich structure. The several factors--high density of biobarcode hpDNA on the surface of AuNPs, multiple electroactive complex molecules intercalated with each hpDNA and dsDNA molecule, and the intercalation binding mode of the electroactive complex with the DNA sandwich structure--contribute to the DNA sensor with highly selective and sensitive sensing properties. The DNA sensor exhibited a detection limit of 1 × 10(-15) M (i.e., 1 fM), the DNA levels in physiological samples, with linearity down to 2 × 10(-15) M. It can differentiate even one single mismatched DNA from the complementary tDNA. This novel biobarcode-based DNA sensing approach should provide a general platform for development of direct, simple, repetitive, sensitive, and selective DNA sensors for various important applications in analytical, environmental, and clinical chemistry.
Assuntos
Sondas de DNA/química , DNA Bacteriano/análise , Técnicas Eletroquímicas/instrumentação , Ouro/química , Helicobacter pylori/isolamento & purificação , Nanopartículas Metálicas/química , Complexos de Coordenação/química , DNA Bacteriano/isolamento & purificação , Técnicas Eletroquímicas/métodos , Eletrodos , Desenho de Equipamento , Infecções por Helicobacter/diagnóstico , Infecções por Helicobacter/microbiologia , Humanos , Ácidos Nucleicos Imobilizados/química , Limite de Detecção , Rutênio/químicaRESUMO
The superconducting magnet with a high magnetic force field can levitate diamagnetic materials. In this study, a specially designed superconducting magnet with large gradient high magnetic field (LGHMF), which provides three apparent gravity levels (µg, 1 g, and 2 g), was used to study its influence on receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation from preosteoclast cell line RAW264.7. The effects of LGHMF on the viability, nitric oxide (NO) production, morphology in RAW264.7 cells were detected by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method, the Griess method, and the immunofluorescence staining, respectively. The changes induced by LGHMF in osteoclast formation, mRNA expression, and bone resorption were determined by tartrate-resistant acid phosphatase staining, semiquantity PCR, and bone resorption test, respectively. The results showed that: 1) LGHMF had no lethal effect on osteoclast precursors but attenuated NO release in RAW264.7 cells. 2) Diamagnetic levitation (µg) enhanced both the formation and bone resorption capacity of osteoclast. Moreover, diamagnetic levitation up-regulated mRNA expression of RANK, Cathepsin K, MMP-9, and NFATc1, while down-regulated RunX2 in comparison with controls. Furthermore, diamagnetic levitation induced obvious morphological alterations in osteoclast, including active cytoplasmic peripheral pseudopodial expansion, formation of pedosome belt, and aggregation of actin ring. 3) Magnetic field produced by LGHMF attenuated osteoclast resorption activity. Collectively, LGHMF with combined effects has multiple effects on osteoclast, which attenuated osteoclast resorption with magnetic field, whereas promoted osteoclast differentiation with diamagnetic levitation. Therefore, these findings indicate that diamagnetic levitation could be used as a novel ground-based microgravity simulator, which facilitates bone cell research of weightlessness condition.