Objective To investigate the isolation and culture of porcine bone marrow mesenchymal stem cell (BMSC) with α-1, 3-galactosyltransferase (GGTA1) gene knockout (GTKO), GTKO/ human CD46 (hCD46) insertion and cytidine monopho-N-acetylneuraminic acid hydroxylase (CMAH)/GGTA1 gene knockout (Neu5GC/Gal), and the protective effect of co-culture with porcine islets on islet cells. Methods Bone marrow was extracted from different transgenic pigs modified with GTKO, GTKO/hCD46 and Neu5GC/Gal. Porcine BMSC were isolated by the whole bone marrow adherent method and then cultured. The morphology of BMSC was observed and the surface markers of BMSC were identified by flow cytometry. Meantime, the multi-directional differentiation induced by BMSC was observed, and the labeling and tracing of BMSC were realized by green fluorescent protein (GFP) transfection. The porcine BMSC transfected with GFP were co-cultured with porcine islet cells. Morphological changes of porcine islet cells were observed, and compared with those in the porcine islet cell alone culture group. Results BMSC derived from pigs were spindle-shaped in vitro, expressing biomarkers of CD29, CD44, CD73, CD90, CD105 and CD166 rather than CD34 and CD45. These cells were able to differentiate into adipocytes, osteoblasts and chondrocytes. Porcine BMSC with GFP transfection could be labeled and traced, which could be stably expressed in the daughter cells after cell division. Porcine BMSC exerted certain protective effect on islet cells. Conclusions GFP-labeled porcine BMSC modified with GTKO, GTKO/hCD46 and Neu5GC/Gal are successfully established, which exert certain protective effect upon islet cells.

Pyroptosis, an inflammatory caspase-dependent programmed cell death, plays a vital role in maintaining tissue homeostasis and activating inflammatory responses. Orthodontic tooth movement (OTM) is an aseptic force-induced inflammatory bone remodeling process mediated by the activation of periodontal ligament (PDL) progenitor cells. However, whether and how force induces PDL progenitor cell pyroptosis, thereby influencing OTM and alveolar bone remodeling remains unknown. In this study, we found that mechanical force induced the expression of pyroptosis-related markers in rat OTM and alveolar bone remodeling process. Blocking or enhancing pyroptosis level could suppress or promote OTM and alveolar bone remodeling respectively. Using Caspase-1^-/- mice, we further demonstrated that the functional role of the force-induced pyroptosis in PDL progenitor cells depended on Caspase-1. Moreover, mechanical force could also induce pyroptosis in human ex-vivo force-treated PDL progenitor cells and in compressive force-loaded PDL progenitor cells in vitro, which influenced osteoclastogenesis. Mechanistically, transient receptor potential subfamily V member 4 signaling was involved in force-induced Caspase-1-dependent pyroptosis in PDL progenitor cells. Overall, this study suggested a novel mechanism contributing to the modulation of osteoclastogenesis and alveolar bone remodeling under mechanical stimuli, indicating a promising approach to accelerate OTM by targeting Caspase-1.
Animals ; Humans ; Mice ; Rats ; Bone Remodeling/physiology* ; Caspase 1 ; Periodontal Ligament ; Pyroptosis ; Tooth Movement Techniques

ObjectiveThe lung mesenchymal stem cells （LMSCs） induced by D-galactose （D-gal） were intervened by Wenfei Huaxian decoction-containing serum to explore the mechanism of Wenfei Huaxian decoction in delaying the senescence of LMSCs through the nicotinamide phosphoribosyltransferase/silent information regulator 1 （NAMPT/SIRT1） signaling pathway. MethodWenfei Huaxian decoction-containing serum was prepared. LMSCs were isolated by gradient density centrifugation， and they were cultured and identified in vitro. The senescence model in vitro was established by stimulating cells via D-gal for 24 h. LMSCs cells were modeled after being treated with different volume fractions （5%， 10%， 20%， 40%， and 80%） of Wenfei Huaxian decoction-containing serum for 24 h， and the cell proliferation level was detected by methyl thiazolyl tetrazolium （MTT） method. The cells were randomly divided into blank serum group， model group， and high， medium， and low dose groups of Wenfei Huaxian decoction-containing serum. Senescence-associated β-galactosidase （SA-β-gal） staining was used to detect the senescence of LMSCs in each group. The content of NAD + was detected by colorimetry. The levels of senescence-associated factors （p16 and p53）， interleukin-6 （IL-6）， and tumor necrosis factor-α （TNF-α） in cell culture supernatant were detected by enzyme-linked immunosorbent assay （ELISA）. Western blot was used to detect the relative expression of senescence-associated proteins and NAMPT/SIRT1 signaling pathway-related proteins. ResultCompared with the blank serum group， the proliferation of LMSCs was significantly inhibited after D-gal stimulation for 24 h （P<0.01）. Compared with the model group， the proliferation of LMSCs could be promoted after intervention with the corresponding Wenfei Huaxian decoction-containing serum （P<0.05， P<0.01）. Compared with the blank serum group， the SA-β-gal staining of LMSCs in the model group after D-gal stimulation was enhanced， and the content of NAD⁺ was increased （P<0.01）. The expression levels of senescence factors p16 and p53， as well as SASP pro-inflammatory factors IL-6 and TNF-α in the cell culture supernatant， were significantly increased （P<0.01）. The expression of senescence-associated proteins p16， p21， and p53 increased （P<0.01）， and the protein expression of NAMPT， SIRT1， peroxisome proliferator-activated receptor γ coactivator 1α （PGC-1α）， and forkhead box family transcription factor O1 （FoxO1） decreased （P<0.01）. Compared with the model group， the SA-β-gal staining of LMSCs in each group of Wenfei Huaxian decoction-containing serum was significantly reduced， and the content of NAD⁺ was decreased （P<0.01）. The senescence factors （p16 and p53） and inflammatory factors （IL-6 and TNF-α） in the cell culture supernatant were significantly decreased （P<0.01）. The expression of senescence-associated proteins （P16， P21， and P53） decreased （P<0.05， P<0.01）. The protein expressions of NAMPT， SIRT1， PGC-1α， and FoxO1 were significantly up-regulated （P<0.05， P<0.01）. ConclusionWenfei Huaxian decoction can alleviate senescence and inflammatory response damage of D-gal-induced LMSCs， and its mechanism may be related to the regulation of the NAMPT/SIRT1 signaling pathway.

Islet transplantation is considered as one of the most effective approach for type 1 diabetes mellitus, although its efficacy is limited by several factors. Anoxia, stress and rejection occurring during the isolation, culturing and transplantation of islets may have impact on the outcome of the islet transplantation. Due to the biological properties such as anti-inflammation, angiogenetic promotion and immune regulation, mesenchymal stem cells (MSCs) are all the way focused by researchers. Additionally, exosome, a derivative of MSC, also plays an import role in regulating anoxia-induced oxidative stress modulation, angiogenetic promotion, and immune regulation. MSC-based islet transplantation may be a useful therapeutic tool in treating type 1 diabetes. Therefore, in this review, the potential effect of MSC prior and posterior to the operation of the islet transplantation, its clinical application as well as its limitations were reviewed, aiming to offer insights into the future application of islet transplantation in treating type 1 diabetes.

ObjectiveTo investigate the possible mechanism of Wenfei Huaxian Decoction （温肺化纤汤） in treatment of pulmonary fibrosis in systemic sclerosis-associated interstitial lung disease （SSc-ILD）. MethodsSixty C3H/He female rats were randomly divided into a control group， a model group， a pirfenidone group， and low-， medium-， and high-dose Wenfei Huaxian Decoction groups. The SSc-ILD model mice was established by subcutaneous injection of bleomycin solution 0.04 mg/d into the back of mice for 28 days in all groups but the control group. After successful modelling， the pirfenidone group was given pirfenidone capsule 300 mg/（kg·d） by gavage， the low-， medium- and high-dose Wenfei Huaxian Decoction groups were given Wenfei Huaxian Decoction 7.81， 15.62， and 31.24 g/（kg·d） by gavage， respectively， and the control group as well as the model group were given normal saline 0.1 ml/10 g by gavage， for a total of 21 days. At the end of the intervention， HE staining and Masson staining were used to observe the pathological changes in the skin and lung tissues； the hydroxyproline content of the skin and lung tissues was detected； the protein expression levels of endoplasmic reticulum stress-related proteins glucose-regulated protein 78 （BIP） and C/EBP homologous protein （CHOP） as well as those of nuclear factor kappa B （NF-κB） pathway p65 were measured by western blot； ELISA was performed to determine the expression levels of interferon gamma （IFN-γ）， interleukin 6 （IL-6） and tumour necrosis factor alpha （TNF-α） in serum of rats. ResultsThe results of HE and Masson staining indicated that compared with the control group， the dermis significantly thickened， the number of collagen fibers significantly enlarged， and the number of inflammatory cells significantly increased in the model group； the lung tissue showed a marked inflammatory cellular response with massive collagen fibre proliferation with inflammatory cell infiltration. Compared with the model group， the skin tissue and lung tissue collagen fibre proliferation significantly reduced and inflammatory cell infiltration reduced in the pirfenidone group and all dose groups of Wenfei Huaxian Decoction， and the effects of pirfenidone group and Wenfei Huaxian Decoction medium- and high-dose groups were basically comparable. Compared with the model group， the content of hydroxyproline in skin and lung tissue， the serum level of IFN-γ， IL-6 and TNF-α， and the expression levels of BIP and CHOP protein in lung tissue increased in model group （P＜0.05）. Compared with model group， the content of hydroxyproline in skin tissue of pirfenidone group， low-and medium-dose Wenfei Huaxian Decoction groups decreased， and the content of hydroxyproline in lung tissue of medium-dose Wenfei Huaxian Decoction group decreased. The serum level of IFN-γ， IL-6， TNF-α and the expression levels of BIP， CHOP and p65 protein in lung tissue of rats in pirfenidone group and high-dose Wenfei Huaxian Decoction group decreased （P＜0.05）. The content of hydroxyproline in lung tissue of medium-dose Wenfei Huaxian Decoction group was significantly lower than that of low-dose and high-dose Wenfei Huaxian Decoction group， and the serum level of IFN-γ， IL-6， TNF-α in low- and medium-dose Wenfei Huaxian Decoction group were higher than those in high-dose Wenfei Huaxian Decoction group. The expression level of BIP protein in high-dose group was significantly lower than that in low- and medium-dose Wenfei Huaxian Decoction groups （P＜0.05）. ConclusionWenfei Huaxian Decoction can improve the skin and lung fibrosis of SSc-ILD rats， which may act through anti-inflammation， inhibition of NF-κB pathway， and then inhibition of endoplasmic reticulum stress， which ultimately blocked the fibrotic process.

Pyroptosis,an inflammatory caspase-dependent programmed cell death,plays a vital role in maintaining tissue homeostasis and activating inflammatory responses.Orthodontic tooth movement(OTM)is an aseptic force-induced inflammatory bone remodeling process mediated by the activation of periodontal ligament(PDL)progenitor cells.However,whether and how force induces PDL progenitor cell pyroptosis,thereby influencing OTM and alveolar bone remodeling remains unknown.In this study,we found that mechanical force induced the expression of pyroptosis-related markers in rat OTM and alveolar bone remodeling process.Blocking or enhancing pyroptosis level could suppress or promote OTM and alveolar bone remodeling respectively.Using Caspase-1-/-mice,we further demonstrated that the functional role of the force-induced pyroptosis in PDL progenitor cells depended on Caspase-1.Moreover,mechanical force could also induce pyroptosis in human ex-vivo force-treated PDL progenitor cells and in compressive force-loaded PDL progenitor cells in vitro,which influenced osteoclastogenesis.Mechanistically,transient receptor potential subfamily V member 4 signaling was involved in force-induced Caspase-1-dependent pyroptosis in PDL progenitor cells.Overall,this study suggested a novel mechanism contributing to the modulation of osteoclastogenesis and alveolar bone remodeling under mechanical stimuli,indicating a promising approach to accelerate OTM by targeting Caspase-1.

Pyroptosis,an inflammatory caspase-dependent programmed cell death,plays a vital role in maintaining tissue homeostasis and activating inflammatory responses.Orthodontic tooth movement(OTM)is an aseptic force-induced inflammatory bone remodeling process mediated by the activation of periodontal ligament(PDL)progenitor cells.However,whether and how force induces PDL progenitor cell pyroptosis,thereby influencing OTM and alveolar bone remodeling remains unknown.In this study,we found that mechanical force induced the expression of pyroptosis-related markers in rat OTM and alveolar bone remodeling process.Blocking or enhancing pyroptosis level could suppress or promote OTM and alveolar bone remodeling respectively.Using Caspase-1-/-mice,we further demonstrated that the functional role of the force-induced pyroptosis in PDL progenitor cells depended on Caspase-1.Moreover,mechanical force could also induce pyroptosis in human ex-vivo force-treated PDL progenitor cells and in compressive force-loaded PDL progenitor cells in vitro,which influenced osteoclastogenesis.Mechanistically,transient receptor potential subfamily V member 4 signaling was involved in force-induced Caspase-1-dependent pyroptosis in PDL progenitor cells.Overall,this study suggested a novel mechanism contributing to the modulation of osteoclastogenesis and alveolar bone remodeling under mechanical stimuli,indicating a promising approach to accelerate OTM by targeting Caspase-1.

Pyroptosis,an inflammatory caspase-dependent programmed cell death,plays a vital role in maintaining tissue homeostasis and activating inflammatory responses.Orthodontic tooth movement(OTM)is an aseptic force-induced inflammatory bone remodeling process mediated by the activation of periodontal ligament(PDL)progenitor cells.However,whether and how force induces PDL progenitor cell pyroptosis,thereby influencing OTM and alveolar bone remodeling remains unknown.In this study,we found that mechanical force induced the expression of pyroptosis-related markers in rat OTM and alveolar bone remodeling process.Blocking or enhancing pyroptosis level could suppress or promote OTM and alveolar bone remodeling respectively.Using Caspase-1-/-mice,we further demonstrated that the functional role of the force-induced pyroptosis in PDL progenitor cells depended on Caspase-1.Moreover,mechanical force could also induce pyroptosis in human ex-vivo force-treated PDL progenitor cells and in compressive force-loaded PDL progenitor cells in vitro,which influenced osteoclastogenesis.Mechanistically,transient receptor potential subfamily V member 4 signaling was involved in force-induced Caspase-1-dependent pyroptosis in PDL progenitor cells.Overall,this study suggested a novel mechanism contributing to the modulation of osteoclastogenesis and alveolar bone remodeling under mechanical stimuli,indicating a promising approach to accelerate OTM by targeting Caspase-1.

Pyroptosis,an inflammatory caspase-dependent programmed cell death,plays a vital role in maintaining tissue homeostasis and activating inflammatory responses.Orthodontic tooth movement(OTM)is an aseptic force-induced inflammatory bone remodeling process mediated by the activation of periodontal ligament(PDL)progenitor cells.However,whether and how force induces PDL progenitor cell pyroptosis,thereby influencing OTM and alveolar bone remodeling remains unknown.In this study,we found that mechanical force induced the expression of pyroptosis-related markers in rat OTM and alveolar bone remodeling process.Blocking or enhancing pyroptosis level could suppress or promote OTM and alveolar bone remodeling respectively.Using Caspase-1-/-mice,we further demonstrated that the functional role of the force-induced pyroptosis in PDL progenitor cells depended on Caspase-1.Moreover,mechanical force could also induce pyroptosis in human ex-vivo force-treated PDL progenitor cells and in compressive force-loaded PDL progenitor cells in vitro,which influenced osteoclastogenesis.Mechanistically,transient receptor potential subfamily V member 4 signaling was involved in force-induced Caspase-1-dependent pyroptosis in PDL progenitor cells.Overall,this study suggested a novel mechanism contributing to the modulation of osteoclastogenesis and alveolar bone remodeling under mechanical stimuli,indicating a promising approach to accelerate OTM by targeting Caspase-1.

Pyroptosis,an inflammatory caspase-dependent programmed cell death,plays a vital role in maintaining tissue homeostasis and activating inflammatory responses.Orthodontic tooth movement(OTM)is an aseptic force-induced inflammatory bone remodeling process mediated by the activation of periodontal ligament(PDL)progenitor cells.However,whether and how force induces PDL progenitor cell pyroptosis,thereby influencing OTM and alveolar bone remodeling remains unknown.In this study,we found that mechanical force induced the expression of pyroptosis-related markers in rat OTM and alveolar bone remodeling process.Blocking or enhancing pyroptosis level could suppress or promote OTM and alveolar bone remodeling respectively.Using Caspase-1-/-mice,we further demonstrated that the functional role of the force-induced pyroptosis in PDL progenitor cells depended on Caspase-1.Moreover,mechanical force could also induce pyroptosis in human ex-vivo force-treated PDL progenitor cells and in compressive force-loaded PDL progenitor cells in vitro,which influenced osteoclastogenesis.Mechanistically,transient receptor potential subfamily V member 4 signaling was involved in force-induced Caspase-1-dependent pyroptosis in PDL progenitor cells.Overall,this study suggested a novel mechanism contributing to the modulation of osteoclastogenesis and alveolar bone remodeling under mechanical stimuli,indicating a promising approach to accelerate OTM by targeting Caspase-1.