sTNFRII-Fc modification protects human UC-MSCs against apoptosis/autophagy induced by TNF-α and enhances their efficacy in alleviating inflammatory arthritis

Abstract Background Tumor necrosis factor (TNF)-α inhibitors represented by Etanercept (a fusion protein containing soluble TNF receptor II (sTNFRII) and the Fc segment of human IgG1) play a pivotal role in Rheumatoid arthritis (RA) treatment. However, long-term use increases the risk of infection and tumors for their systemic inhibition of TNF-α, which disrupts the regular physiological function of this molecular. Mesenchymal stem cells (MSCs)-based delivery system provides new options for RA treatment with their “homing” and immune-regulation capacities, whereas inflammatory environment (especially TNF-α) is not conducive to MSCs' therapeutic effects by inducing apoptosis/autophagy. Here, we constructed a strain of sTNFRII-Fc-expressing MSCs (sTNFRII-MSC), aiming to offset the deficiency of those two interventions. Methods Constructed sTNFRII-Fc lentiviral vector was used to infect human umbilical cord-derived MSCs, and sTNFRII-MSC stable cell line was generated by monoclonal cultivation. In vitro and vivo characteristics of sTNFRII-MSC were assessed by coculture assay and an acute inflammatory model in NOD/SCID mice. The sTNFRII-MSC were transplanted into CIA model, pathological and immunological indicators were detected to evaluate the therapeutic effects of sTNFRII-MSC. The distribution of sTNFRII-MSC was determined by immunofluorescence assay. Apoptosis and autophagy were analyzed by flow cytometry, western blot and immunofluorescence. Results sTNFRII-Fc secreted by sTNFRII-MSC present biological activity both in vitro and vivo. sTNFRII-MSC transplantation effectively alleviates mice collagen-induced arthritis (CIA) via migrating to affected area, protecting articular cartilage destruction, modulating immune balance and sTNFRII-MSC showed prolonged internal retention via resisting apoptosis/autophagy induced by TNF-α. Conclusion sTNFRII-Fc modification protects MSCs against apoptosis/autophagy induced by TNF-α, in addition to releasing sTNFRII-Fc neutralizing TNF-α to block relevant immune-inflammation cascade, and thus exert better therapeutic effects in alleviating inflammatory arthritis.

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sTNFRII-Fc modification protects human UC-MSCs against apoptosis/autophagy induced by TNF-α and enhances their efficacy in alleviating inflammatory arthritis

10.21203/rs.3.rs-695405/v1 ◽

2021 ◽

Author(s):

Yingjie Zhao ◽

Xuezhi Yang ◽

Siyu Li ◽

Bingjie Zhang ◽

Susu Li ◽

...

Keyword(s):

Inflammatory Arthritis ◽

Lentiviral Vector ◽

Cartilage Destruction ◽

Stable Cell Line ◽

Human Umbilical Cord ◽

Therapeutic Effects ◽

Immune Balance ◽

Tnf Α ◽

Cultivation In Vitro

Abstract Background Tumor necrosis factor (TNF)-α inhibitors represented by etanercept (a fusion protein containing soluble TNF receptor Ⅱ (sTNFRⅡ) and the Fc segment of human IgG1) play a pivotal role in Rheumatoid arthritis (RA) treatment. However, long-term use increases the risk of infection and tumors for their systemic inhibition of TNF-α, which disrupts the regular physiological function of this molecular. Mesenchymal stem cells (MSCs)-based delivery system provides new options for RA treatment with their “homing” and immune-regulation capacities, whereas inflammatory environment (especially TNF-α) is not conducive to MSCs' therapeutic effects by inducing apoptosis/autophagy. Here, we constructed a strain of sTNFRⅡ-Fc-expressing MSCs (sTNFRⅡ-MSC), aiming to offset the deficiency of those two interventions. Methods Constructed sTNFRII-Fc lentiviral vector was used to infect human umbilical cord-derived MSCs, and sTNFRII-MSC stable cell line was generated by monoclonal cultivation. In vitro and vivo characteristics of sTNFRII-MSC were assessed by coculture assay and an acute inflammatory model in NOD/SCID mice. The sTNFRII-MSC were transplanted into CIA model, pathological and immunological indicators were detected to evaluate the therapeutic effects of sTNFRII-MSC. The distribution of sTNFRII-MSC was determined by immunofluorescence assay. Apoptosis and autophagy was analyzed by flow cytometry, western blot and immunofluorescence. Results sTNFRⅡ-Fc secreted by sTNFRⅡ-MSC present biological activity both in vitro and vivo. sTNFRⅡ-MSC transplantation effectively alleviates mice collagen-induced arthritis (CIA) via migrating to affected area, protecting articular cartilage destruction, modulating immune balance and sTNFRⅡ-MSC showed prolonged internal retention via resisting apoptosis/autophagy induced by TNF-α. Conclusion sTNFRⅡ-Fc modification protects MSCs against apoptosis/autophagy induced by TNF-α, in addition to releasing sTNFRⅡ-Fc neutralizing TNF-α to block relevant immune-inflammation cascade, and thus exert better therapeutic effects in alleviating inflammatory arthritis.

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Alleviation of Oxidative Stress in Dental Pulp Cells Following 4-Hexylresorcinol Administration in a Rat Model

Applied Sciences ◽

10.3390/app11083637 ◽

2021 ◽

Vol 11 (8) ◽

pp. 3637

Author(s):

Jun-Ho Chang ◽

Dae-Won Kim ◽

Seong-Gon Kim ◽

Tae-Woo Kim

Keyword(s):

Oxidative Stress ◽

Dental Pulp ◽

Therapeutic Effects ◽

Protective Effects ◽

Mandibular Incisor ◽

Tnf Α ◽

Total Antioxidant ◽

Pulp Cells ◽

Pre Treatment

Damaged dental pulp undergoes oxidative stress and 4-hexylresorcinol (4HR) is a well-known antioxidant. In this study, we aimed to evaluate the therapeutic effects of a 4HR ointment on damaged dental pulp. Pulp cells from rat mandibular incisor were cultured and treated with 4HR or resveratrol (1–100 μM). These treatments (10–100 μM) exerted a protective effect during subsequent hydrogen peroxide treatments. The total antioxidant capacity and glutathione peroxidase activity were significantly increased following 4HR or resveratrol treatment (p < 0.05), while the expression levels of TNF-α and IL1β were decreased following the exposure to 4HR pre-treatment in an in vitro model. Additionally, the application of 4HR ointment in an exposed dental pulp model significantly reduced the expression of TNF-α and IL1β (p < 0.05). Conclusively, 4HR exerted protective effects against oxidative stress in dental pulp tissues through downregulating TNF-α and IL1β.

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Skin fibroblasts' alteration after Acupuncture on LI11 in Rabbits with bacteria endotoxin induced fever

Acupuncture & Electro-Therapeutics Research ◽

10.3727/036012921x16112663844932 ◽

2021 ◽

Vol 45 (2) ◽

pp. 125-136

Author(s):

Wang Fan ◽

Fan Shendong ◽

Cui Guangwei ◽

Cheng Huaijin ◽

Shi She ◽

...

Keyword(s):

Morphological Changes ◽

Therapeutic Effects ◽

Atp Content ◽

Model Group ◽

Tnf Α ◽

Significant Difference ◽

Group A ◽

Subcutaneous Connective Tissue ◽

Needle Retaining

Objectives: To study the mechanism of acupuncture manipulation (AM) based on the secretion function and morphological variation of fibroblasts in acupoint region.<br/> Methods: 40 rabbits were randomly divided to normal group (N), model group (M), needle retaining group (A), and acupuncture manipulation group (AM), each group consist of 10 rabbits. The animal model of Rabbits with Bacterium Endotoxin Induced Fever (BEIF) was established by intravenous injection of bacterial endotoxin. Groups A and AM were treated with 'Qu Chi'(LI11) acupuncture after modeling, and acupuncture manipulations were conducted in group AM. The serum heat factors IL-1β, IL-6, and TNF-α content were assayed conducted in group AM. The serum heat factors IL-1β, IL-6, and TNF-α content were assayed by Elisa after acupuncturet; the morphological changes of fibroblasts in acupoint area were observed by Vimentin staining; the fibroblasts of subcutaneous connective tissue in acupoint area were isolated and cultured in vitro, and the contents of PEG2, NO and ATP in supernatant were assayed.<br/> Results: There was no significant difference in fibroblasts cytomorphology among groups M, A, and N. Fibroblasts in group AM were stretched and aligned in almost one direction. Comparing with group N, the content of serum IL-1β, IL-6, and TNF-α was significantly higher in group M, along with higher NO and ATP content in the cell culture supernant; Comparing with group M, content of IL-6 and TNF-α was lower in group A, content of serum IL-1β, IL-6 and TNF-α was higher in group AM, along with higher PEG2, NO and ATP content in both groups, while group AM demonstrated more significant changes in the above indicators than group A.<br/> Conclusion: Acupuncture had therapeutic effects on BEIF rabbits, the application of acupuncture manipulation can further improve its therapeutic effects. The mechanism may be related to the influence on fibroblasts cytomorphology in acupoint region and facilitation of fibroblasts-derived PEG2, NO, and ATP, which together promote the acupoint vitality.

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SARS-CoV-2 Spike Protein S1-Mediated Endothelial Injury and Pro-Inflammatory State Is Amplified by Dihydrotestosterone and Prevented by Mineralocorticoid Antagonism

Viruses ◽

10.3390/v13112209 ◽

2021 ◽

Vol 13 (11) ◽

pp. 2209

Author(s):

Nitin Kumar ◽

Yu Zuo ◽

Srilakshmi Yalavarthi ◽

Kristina L. Hunker ◽

Jason S. Knight ◽

...

Keyword(s):

Endothelial Cells ◽

Endothelial Activation ◽

Endothelial Injury ◽

Spike Protein ◽

Severe Illness ◽

Therapeutic Effects ◽

Vascular Effects ◽

Adjunct Treatment ◽

Tnf Α

Men are disproportionately affected by the coronavirus disease-2019 (COVID-19), and face higher odds of severe illness and death compared to women. The vascular effects of androgen signaling and inflammatory cytokines in severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-mediated endothelial injury are not defined. We determined the effects of SARS-CoV-2 spike protein-mediated endothelial injury under conditions of exposure to androgen dihydrotestosterone (DHT) and tumor necrosis factor-a (TNF-α) and tested potentially therapeutic effects of mineralocorticoid receptor antagonism by spironolactone. Circulating endothelial injury markers VCAM-1 and E-selectin were measured in men and women diagnosed with COVID-19. Exposure of endothelial cells (ECs) in vitro to DHT exacerbated spike protein S1-mediated endothelial injury transcripts for the cell adhesion molecules E-selectin, VCAM-1 and ICAM-1 and anti-fibrinolytic PAI-1 (p < 0.05), and increased THP-1 monocyte adhesion to ECs (p = 0.032). Spironolactone dramatically reduced DHT+S1-induced endothelial activation. TNF-α exacerbated S1-induced EC activation, which was abrogated by pretreatment with spironolactone. Analysis from patients hospitalized with COVID-19 showed concordant higher circulating VCAM-1 and E-Selectin levels in men, compared to women. A beneficial effect of the FDA-approved drug spironolactone was observed on endothelial cells in vitro, supporting a rationale for further evaluation of mineralocorticoid antagonism as an adjunct treatment in COVID-19.

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Exosomes derived from mesenchymal stem cells inhibit catabolism in human chondrocytes by activating autophagy via inhibition of the NF-κB pathway

10.21203/rs.3.rs-83574/v1 ◽

2020 ◽

Author(s):

Jian-Fa Wang ◽

Zhi Zhu ◽

Lei Sun ◽

Shi-kun Shao ◽

Bao-dong Ma ◽

...

Keyword(s):

Electron Microscopy ◽

Primary Cultures ◽

Human Chondrocytes ◽

Inflammatory Factors ◽

Human Umbilical Cord ◽

Adult Males ◽

New Therapeutic Approaches ◽

Inflammatory Conditions ◽

Tnf Α

Abstract Objective: We aimed to determine the significance of MSC-derived exosomes (MSC-Exos) in chondrocyte autophagy under normal and inflammatory conditions.Design: Human umbilical cord-derived MSCs (hMSCs) were cultured in vitro. hMSC-Exos( EX） were isolated by an ultracentrifugation method. Transmission electron microscopy and western analysis were used to identify exosomes. Human chondrocytes were extracted from five adult males with OA undergoing total knee arthroplasty. Primary cultures of chondrocytes from OA patients were stimulated with 50 ng/ml tumor necrosis factor-α (TNF-α) in the presence or absence of hMSC-Exos. Autophagy levels were determined based on expression of autophagic marker LC3, StubRFP-SensGFP-LC3 analysis, and electron microscopy. Catabolic gene and chemokine expression were evaluated using quantitative PCR. The NF-κB inhibitor NS398 was used to analyze the role of the NF-κB pathway in autophagic activation.Results: hMSC-Exos increased LC3-II levels as well as autophagosome number in chondrocytes. hMSC-Exos inhibited TNF-α–induced expression of MMP-3, -9, and -13; ADAMTS5; CCL-2 and -5; and CXCL1. NF-κB inhibition activated autophagy in TNF-α–treated chondrocytes. These results indicate that hMSC-Exos might suppress the levels of catabolic and inflammatory factors in chondrocytes by promoting autophagy via NF-κB pathway inhibition.Conclusions: Our data support the interest in hMSC-Exos to develop new therapeutic approaches for joint conditions.

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Human Umbilical Mesenchymal Stromal Cells Mixed with Hyaluronan Transplantation Decreased Cartilage Destruction in a Rabbit Osteoarthritis Model

Stem Cells International ◽

10.1155/2021/2989054 ◽

2021 ◽

Vol 2021 ◽

pp. 1-12

Author(s):

Yu-Hsun Chang ◽

Dah-Ching Ding ◽

Kun-Chi Wu

Keyword(s):

Mesenchymal Stromal Cells ◽

Stromal Cells ◽

Rabbit Model ◽

Human Leukocyte ◽

Type Ii Collagen ◽

Cartilage Destruction ◽

Human Umbilical Cord ◽

Safranin O

Osteoarthritis (OA), the most common type of arthritis, causes pain in joints and disability. Due to the absence of ideal effective medication, stem cell transplantation emerges as a new hope for OA therapy. This study is aimed at evaluating the capability of human umbilical cord mesenchymal stromal cells (HUCMSCs) mixed with hyaluronan (HA) to treat osteoarthritis in a rabbit model. Differentiation capability of HUCMSCs, magnetic resonance image examination, and immunohistochemistry of the cartilage after transplantation of HUCMSCs mixed with HA in a rabbit OA model were explored. HUCMSCs exhibited typical mesenchymal stromal cell (MSC) characteristics, including spindle-shaped morphology, surface marker expressions (positive for human leukocyte antigen- (HLA-) ABC, CD44, CD73, CD90, and CD105; negative for HLA-DR, CD34, and CD45), and trilineage differentiation (chondrogenesis, adipogenesis, and osteogenesis). The gene expression of SOX9, type II collagen, and aggrecan in the HUCMSC-derived chondrocytes mixed with HA was increased after in vitro chondrogenesis compared with HUCMSCs. A gross and histological significant improvement in hyaline cartilage destruction after HUCMSCs mixed with HA was noted in the animal model compared to the OA knees. The International Cartilage Repair Society histological score and Safranin O staining were significantly higher for the treated knees than the control knees ( p < 0.05 ). Moreover, the expression of MMP13 was significantly decreased in the treated knees than in the OA knees. In conclusion, HUCMSCs mixed with HA in vitro and in vivo might attenuate the cartilage destruction in osteoarthritis. Our study provided evidence for future clinical trials.

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Human Umbilical Cord-Derived Mesenchymal Stem Cells Alleviate Acute Lung Injury Caused by Severe Burn via Secreting TSG-6 and Inhibiting Inflammatory Response

10.21203/rs.3.rs-127492/v2 ◽

2021 ◽

Author(s):

Xiaohong Hu ◽

Lingying Liu ◽

Yu Wang ◽

Zhongyuan Li ◽

Yonghui Yu ◽

...

Keyword(s):

Lung Injury ◽

Pulmonary Function ◽

Human Umbilical Cord ◽

Ct Scanner ◽

Severe Burn ◽

Positive Effects ◽

Tnf Α ◽

Anti Inflammatory Cytokine

Abstract Objectives To investigate whether hUC-MSCs attenuated severe burn-induced ALI and the effects were based on TSG-6 secreted from hUC-MSCs. Method Rat model was established and evaluated as follows：Anires2005 animal pulmonary function tester for pulmonary function; micro-CT scanner for lung imaging manifestations; cytokine expression was measured by ELISA assay, and both inflammatory cell infiltration and lung injury were assessed by immunohistochemistry assay. Results In vitro, TSG-6 levels in serum from the burn group were significantly increased than that of the sham group. In vivo, TSG-6 levels of lung tissues and serum in the burn+ hUC-MSCs group were significantly increased than those of that in the burn group. Higher parameters of airway resistance（Ri, Re, etc）were markedly decreased, and the disordered lung texture and funicular density shadows were significantly improved after hUC-MSCs administration. Both in lung tissues and serum, increased levels of proinflammatory cytokines（TNF-α, IL-1β, IL-6）were remarkably decreased, but anti-inflammatory cytokine IL-10 increased after hUC-MSCs administration (p<0.05). These significant positive effects after hUC-MSCs transplantation did not occur in the Burn+siTSG-6 group. Conclusion Intra-tracheal implantation of hUC-MSCs has been an effective treatment for severe burn-induced ALI via promoting TSG-6 secretion and inhibiting inflammatory reaction in lung tissue.

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Exosomes derived from human placental MSCs carrying miR-4450 inhibitor alleviate intervertebral disc degeneration and ameliorate gait disturbances via up-regulation of ZNF121

10.21203/rs.3.rs-28561/v1 ◽

2020 ◽

Author(s):

Qiling Yuan ◽

Xinyi Wang ◽

Liang Liu ◽

Yongsong Cai ◽

Xiaoming Zhao ◽

...

Keyword(s):

Intervertebral Disc ◽

Disc Degeneration ◽

Intervertebral Disc Degeneration ◽

Therapeutic Effects ◽

Nucleus Pulposus Cells ◽

Gait Abnormality ◽

Tnf Α ◽

Potential Therapeutic Role

Abstract Background Exosomes derived from mesenchymal stem cells (MSCs) have emerged as novel drug and gene delivery tools. Current study aimed to elucidate the potential therapeutic role of human placental MSC (hPLMSC)-derived exosomes carrying antagomiR-4450 (EXO-antagomiR-4450) in intervertebral disc degeneration (IDD) progression. Methods Initially, the differentially expressed miRNAs related to IDD were identified by microarray analysis which provided data predicting the interaction between miR-4450 and ZNF121 in IDD. Next, miR-4450 and ZNF121 were elevated or silenced to determine their effects on the damage of NPCs treated with TNF-α. The therapeutic effects of EXO-antagomiR-4450 on nucleus pulposus cells (NPCs) were verified both in vitro and in vivo, especially gait analysis and fluorescent molecular tomopraphy were used in live IDD mice. Results Our results revealed that miR-4450 was highly expressed, while ZNF121 was poorly expressed in IDD patients and NPCs treated with TNF-α. Furthermore, miR-4450 was identified to specifically target ZNF121. Additionally, the inhibition of miR-4450 exerted an alleviatory effect on the inflammation, apoptosis and damage of the NPCs by up-regulating ZNF121. Moreover, EXO-antagomiR-4450 retarded damage of NPCs in vitro, alleviated IDD damage and ameliorated gait abnormality in vivo. Conclusion hPLMSC-derived exosomes could be a feasible nanovehicle to deliver inhibitory oligonucleotides like antagomiR-4450 in IDD.

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Efficient Intracytoplasmic Labeling of Human Umbilical Cord Blood Mesenchymal Stromal Cells with Ferumoxides

Cell Transplantation ◽

10.3727/000000007783465271 ◽

2007 ◽

Vol 16 (8) ◽

pp. 849-857 ◽

Cited By ~ 23

Author(s):

Jae Kwon Lee ◽

Man Kyoung Lee ◽

Hye Jin Jin ◽

Dal-Soo Kim ◽

Yoon Sun Yang ◽

...

Keyword(s):

Mesenchymal Stromal Cells ◽

Stromal Cells ◽

Superparamagnetic Iron Oxide ◽

Cell Labeling ◽

Human Umbilical Cord Blood ◽

Human Umbilical Cord ◽

Therapeutic Effects ◽

Noninvasive Technique

Mesenchymal stromal cells (MSCs) are multipotent cells found in several adult tissues; they have the capacity to differentiate into mesodermal, ectodermal, and endodermal tissues in vitro. There have been several reports that MSCs have therapeutic effects in a variety of diseases. Therefore, using a cell labeling technique, monitoring their temporal and spatial migration in vivo, would be useful in the clinical setting. Magnetic resonance imaging (MRI)—tracking of superparamagnetic iron oxide (SPIO)-labeled cells—is a noninvasive technique for determining the location and migration of transplanted cells. In the present study, we evaluated the influence and toxicity of SPIO (ferumoxides) labeling on multiple differentiated MSCs. To evaluate the influence and toxicity of ferumoxides labeling on differentiation of MSCs, a variety of concentrations of ferumoxides were used for labeling MSCs. We found that the cytoplasm of adherent cells was effectively labeled at low concentrations of ferumoxides. Compared with unlabeled controls, the ferumoxides-labeled MSCs exhibited a similar proliferation rate and apoptotic progression. The labeled MSCs differentiated into osteoblasts and adipocytes in an identical fashion as the unlabeled cells. However, chondrogenesis and neurogenesis were inhibited at high concentrations of ferumoxides. Our results suggest the effective concentration for ferumoxides use in tracking MSCs.

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Profile of host cell responses to exposure to stressed bacteria in planktonic; dislodged, and intact biofilm mode

Brazilian Dental Journal ◽

10.1590/0103-6440202104503 ◽

2021 ◽

Vol 32 (3) ◽

pp. 10-20

Author(s):

Rafaela Fernandes Zancan ◽

José Burgos Ponce ◽

Thiago José Dionisio ◽

Rodrigo Cardoso de Oliveira ◽

Rafaela Alves da Silva ◽

...

Keyword(s):

Antimicrobial Activity ◽

Enterococcus Faecalis ◽

Inflammatory Cytokines ◽

Human Peripheral Blood ◽

Negative Control ◽

Therapeutic Effects ◽

Tnf Α ◽

Host Defense Response ◽

Post Hoc

Abstract The host defense response to microbial challenge emerging from the root canal system leads to apical periodontitis. The aim of this study was to evaluate the expression of inflammatory cytokines and Nitric Oxide (NO) by macrophages after interaction with Enterococcus faecalis in the: plankton and dislodged biofilm mode; intact biofilm mode stimulated by calcium hydroxide (CH), CH and chlorhexidine (CHX) or Triple Antibiotic Paste (TAP). For this purpose, culture of macrophages from monocytes in human peripheral blood (N=8) were exposed to the different modes of bacteria for 24 hours. Subsequently, the cytokines, such as, Tumor Necrotic Factor- alfa (TNF-α), interleukin (IL)-1β, IL-6, IL-10; and NO were quantified by Luminex xMAP and Greiss reaction, respectively. In addition to the potential therapeutic effects of the intracanal medication, their antimicrobial activity against Enterococcus faecalis biofilm were also tested in vitro by confocal microscopy. The experiments` data were analyzed by the Kruskal-Wallis test with the Dunn post hoc test (α < 0.05). Bacteria in dislodged biofilm mode were shown to be more aggressive to the immune system than bacteria in plankton mode and negative control, inducing greater expression of NO and TNF-α. Relative to bacteria in intact biofilm mode, the weakest antimicrobial activity occurred in Group CH. In Groups CH/CHX and TAP the percentage of dead bacteria was significantly increased to the same extent. Interestingly, the biofilm itself did not induce the release of pro-inflammatory cytokines - except for NO - while the biofilm treated with TAP and CH based pastes enhanced the levels of IL-6 and TNF-α; and IL-1 β, respectively. In contrast, the levels of a potent anti-inflammatory (IL-10) were increased in Group TAP.

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