Guijuan Feng

TNF-α triggers osteogenic differentiation of human dental pulp stem cells via the NF-κB signalling pathway.

Dental pulp stem cells (DPSCs) are a type of mesenchymal stem cells (MSCs) characterised by self‐renewal and multi‐lineage differentiation, including chondrocytes, adipocytes, neural cells and osteoblasts, which make it an attractive choice for tissue engineering purposes. Tumour necrosis factor α (TNF‐α) had the positive effect on the mineralisation of bone marrow MSCs and stromal cells derived from human adipose tissue. However, the effect of TNF‐α on DPSCs is unclear. We found that TNF‐α activated the NF‐κB pathway during the osteogenic differentiation of DPSCs. TNF‐α also increased mineralisation and the expression of bone morphogenetic protein 2 (BMP2), alkaline phosphatase (ALP), runt‐related transcription factor 2 (RUNX2) and collagen type I (COL I) during this process. PDTC, an NF‐κB inhibitor, blocked the osteogenic differentiation induced by TNF‐α. No effect of TNF‐α on proliferation of DPSCs or cell cycle was detected. In summary, TNF‐α promotes mineralisation and mineralisation‐related gene expression through the NF‐κB signalling pathway in DPSCs, which may provide a foundation for autologous transplantation of DPSCs.

Insulin‐like growth factor 1 can promote proliferation and osteogenic differentiation of human dental pulp stem cells via mTOR pathway

Insulin‐like growth factor 1 (IGF‐1) is a multifunctional peptide that can enhance osteogenic differentiation of bone marrow mesenchymal stem cells (BMMSCs). However, it remains unclear whether IGF‐1 can promote osteogenic differentiation of human dental pulp stem cells (DPSCs). In our study, DPSCs were isolated from the impacted third molars, and treated with IGF‐1. Osteogenic differentiation abilities were investigated. We found that IGF‐1 activated the mTOR signaling pathway during osteogenic differentiation of DPSCs. IGF‐1 also increased the expression of runt‐related transcription factor 2 (RUNX2), osteocalcin (OCN), osterix (OSX) and collagen type I (COL I) during this process. Rapamycin, an mTOR inhibitor, blocked osteogenic differentiation induced by IGF‐1. Meanwhile, CCK‐8 assay and flow cytometry results demonstrated that 10–200 ng/mL IGF‐1 could enhance proliferation ability of DPSCs and 100 ng/mL was the optimal concentration. In summary, IGF‐1 could promote proliferation and osteogenic differentiation of DPSCs via mTOR pathways, which might have clinical implications for osteoporosis.

The Correlations of Disease Activity, Socioeconomic Status, Quality of Life, and Depression/Anxiety in Chinese Patients with Systemic Lupus Erythematosus

The prevalence of psychological problems is frequent in systemic lupus erythematosus (SLE) patients and appears to be increasing. The current study investigated the relationship among disease parameters, quality of life, and the psychological status in Chinese patients with SLE. A self-report survey design was administered to 170 SLE patients and 210 healthy individuals using the Self-Rating Anxiety Scale, the Self-Rating Depression Scale, and the Short Form 36 health survey (SF-36). Our results showed that 20.3% SLE patients had anxiety, and 32.9% had depression, which were significantly higher than the control group (7.1%, 14.3%, resp.). And there were significant correlations among socioeconomic status (SES), disease activity, and anxiety/depression in SLE patients. Meanwhile, SF-36 analysis results revealed that VT, PF, and RP scales were the most powerful predictors of anxiety of SLE patients, and SLEDAI, VT, PF, SF, and RE domains were significantly accounted for anxiety. In summary, there were significant relationships among disease parameters, quality of life, and anxiety/depression in Chinese SLE patients. Therefore, it is necessary to have psychiatric and psychological evaluations and formulate an integrated approach for managing mental health in Chinese lupus patients, especially those who have high disease activity, low SES, and poor quality of life.

Human Umbilical Cord-Derived Mesenchymal Stem Cells Downregulate Inflammatory Responses by Shifting the Treg/Th17 Profile in Experimental Colitis

Background/Aims: The aim of this study was to evaluate the effect and mechanisms of human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) on immune responses in murine colitis. Methods: Mice with dextran sulfate sodium (DSS)-induced colitis were injected intraperitoneally with hUC-MSCs or human bone marrow-derived MSCs. The cytokine levels from lamina propria mononuclear cells (LPMCs) and colon tissue were measured using ELISA. Treg and Th17 cells were analyzed using flow cytometry. The proliferation of LPMCs was assessed using Cell Counting Kit-8. Results: hUC-MSCs ameliorate DSS-induced colitis via the downregulation of colon inflammatory responses. Furthermore, hUC-MSCs adjusted modulation of Treg/Th17 cells in the spleen and mesenteric lymph nodes. hUC-MSCs also inhibited LPMCs in vitro.Conclusion: hUC-MSCs may be an alternative source of stem cells and are worthy of study in long-term clinical trials.

RBM5 and p53 expression after rat spinal cord injury: Implications for neuronal apoptosis

RBM5 (RNA-binding motif protein 5), a nuclear RNA binding protein, is known to trigger apoptosis and induce cell cycle arrest by regulating the activity of the tumor suppressor protein p53. However, its expression and function in spinal cord injury (SCI) are still unknown. To investigate whether RBM5 is involved in central nervous system injury and repair, we performed an acute SCI model in adult rats in this study. Our results showed RBM5 was unregulated significantly after SCI, which was accompanied with an increase in the levels of apoptotic proteins such as p53, Bax, and active caspase-3. Immunofluorescent labeling also showed that traumatic SCI induced RBM5 location changes and co-localization with active caspase-3 in neurons. To further probe the role of RBM5, a neuronal cell line PC12 was employed to establish an apoptotic model. Knockdown of RBM5 apparently decreased the level of p53 as well as active caspase-3, demonstrating its pro-apoptotic role in neurons by regulating expressions of p53 and caspase-3. Taken together, our findings indicate that RBM5 promotes neuronal apoptosis through modulating p53 signaling pathway following SCI.

Repeated lipopolysaccharide stimulation promotes cellular senescence in human dental pulp stem cells (DPSCs)

Dental pulp stem cells (DPSCs) are a type of mesenchymal stem cell (MSC) characterized by multi-lineage differentiation making it an attractive choice for tissue regeneration. However, before DPSCs can be used for cell-based therapy, we have to understand their biological properties in response to intrinsic and extrinsic stimuli such as lipopolysaccharide (LPS). DPSCs were therefore stimulated with LPS and senescence was evaluated by senescence-associated β-galactosidase (SA-β-gal) staining, with cell number and cell-cycle arrest being examined by BrdU assay and flow cytometry, respectively. The morphology of DPSCs was characterized by their flat shape, increased size and increased SA-β-gal activity after repeated stimulation (3 or 6 times) with LPS. Reactive oxygen species (ROS) staining showed that the number of ROS-stained cells and the DCFH fluorescent level were higher in the LPS-treated DPSCs compared with those in the untreated DPSCs. Protein and mRNA expression levels of γ-H2A.X and p16INK4A were also increased in DPSCs with repeated LPS stimulation. We found that the LPS bound with Toll-like receptor 4 (TLR4) and that TLR4 signaling accounted for p16INK4A expression. Further results indicated that the senescence of DPSCs stimulated repeatedly with LPS was reversed by p16INK4A short interfering RNA. The DNA damage response and p16INK4A pathways might be the main mediators of DPSC senescence induced by repeated LPS stimulation. Thus, DPSCs tend to undergo senescence after repeated activation, implying that DPSC senescence starts after many inflammatory challenges. Ultimately, these findings should lead to a better understanding of DPSC-based clinical therapy.

p16INK4A mediates age-related changes in mesenchymal stem cells derived from human dental pulp through the DNA damage and stress response

Mesenchymal stem cells derived from human dental pulp (DP-MSCs) are characterized by self-renewal and multi-lineage differentiation, which play important roles in regenerative medicine. Autologous transfers, as non-immunogenic, constitute the safest approach in cellular transplantations. However, their use may be limited by age-related changes. In the study, we compared DP-MSCs isolated from human in five age groups: 5-12 y, 12-20 y, 20-35 y, 35-50 y, and >50 y. We tested the effect of age on proliferation, differentiation, senescence-associated β-galactosidase (SA-β-gal), cell cycle and programmed cell death. DP-MSCs showed characteristics of senescence as a function of age. Meanwhile, the expression of p16(INK4A) and γ-H2A.X significantly increased with age, whereas heat shock protein 60 (HSP60) was decreased in the senescent DP-MSCs. Reactive oxygen species (ROS) staining showed the number of ROS-stained cells and the DCFH fluorescent level were higher in the aged group. Further we examined the senescence of DP-MSCs after modulating p16(INK4A) signaling. The results indicated the dysfunction of DP-MSCs was reversed by p16(INK4A) siRNA. In summary, our study indicated p16(INK4A) pathway may play a critical role in DP-MSCs age-related changes and the DNA damage response (DDR) and stress response may be the main mediators of DP-MSCs senescence induced by excessive activation of p16(INK4A) signaling.

Roles of Wnt/β-catenin signaling in retinal neuron-like differentiation of bone marrow mesenchymal stem cells from nonobese diabetic mice.

Recent studies have shown that mesenchymal stem cells (MSCs) are expected to become promising therapeutic agents for the treatment of diabetic retinopathy (DR); moreover, we previously demonstrated that bone marrow (BM)-MSCs from nonobese diabetic (NOD) mice (an ideal DR model) had abnormal migration and adhesion. So, we hypothesized that NOD-MSCs also have abnormal retinal neuron-like differentiation potential. MSCs were cultured with brain-derived neurotrophic factor, nerve growth factor, and basic fibroblast growth factor. Western blot analysis and immunofluorescence both showed that the level of retinal neuron-like markers, such as glial fibrillary acidic protein, neuron-specific nuclear protein, tyrosine hydroxylase, Thy-1, glutamine synthetase, and rhodopsin was lower in NOD-MSCs than in imprinting control region MSCs. Furthermore, we explored the precise mechanisms controlling this change in NOD-MSCs. The expression levels of some important member proteins in Wnt/β-catenin signaling were determined and suggested the downregulation of Wnt/β-catenin signaling with retinal neuron-like differentiation of NOD-MSCs. Incubation of NOD-MSCs in medium supplemented with human recombinant Wnt1 resulted in a significant upregulation of retinal neuron-like markers, and the effects of Wnt1 were dose-dependent. Taken together, our study indicated that the inhibition of Wnt/β-catenin signaling in NOD-MSCs after induction could contribute to the abnormal retinal neuron-like differentiation. These data provide important preclinical references supporting the basis for further development of autologous MSC-based therapies for DR.Recent studies have shown that mesenchymal stem cells (MSCs) are expected to become promising therapeutic agents for the treatment of diabetic retinopathy (DR); moreover, we previously demonstrated that bone marrow (BM)-MSCs from nonobese diabetic (NOD) mice (an ideal DR model) had abnormal migration and adhesion. So, we hypothesized that NOD-MSCs also have abnormal retinal neuron-like differentiation potential. MSCs were cultured with brain-derived neurotrophic factor, nerve growth factor, and basic fibroblast growth factor. Western blot analysis and immunofluorescence both showed that the level of retinal neuron-like markers, such as glial fibrillary acidic protein, neuron-specific nuclear protein, tyrosine hydroxylase, Thy-1, glutamine synthetase, and rhodopsin was lower in NOD-MSCs than in imprinting control region MSCs. Furthermore, we explored the precise mechanisms controlling this change in NOD-MSCs. The expression levels of some important member proteins in Wnt/β-catenin signaling were determined and suggested the downregulation of Wnt/β-catenin signaling with retinal neuron-like differentiation of NOD-MSCs. Incubation of NOD-MSCs in medium supplemented with human recombinant Wnt1 resulted in a significant upregulation of retinal neuron-like markers, and the effects of Wnt1 were dose-dependent. Taken together, our study indicated that the inhibition of Wnt/β-catenin signaling in NOD-MSCs after induction could contribute to the abnormal retinal neuron-like differentiation. These data provide important preclinical references supporting the basis for further development of autologous MSC-based therapies for DR.

JAB1 accelerates odontogenic differentiation of dental pulp stem cells

Jun activation domain-binding protein 1 (JAB1) is a multifunctional protein that participates in the control of cell proliferation and the stability of multiple proteins. JAB1 regulates several key proteins, and thereby produces varied effects on cell cycle progression, genome stability and cell survival. Some studies have shown that the loss of JAB1 in osteochondral progenitor cells severely impairs embryonic limb development in mice. However, the biological significance of JAB1 activity in the odontogenic differentiation of dental pulp stem cells (DPSCs) remains unclear. This study aimed to determine the role of JAB1, a key player in tooth development, in reparative dentin formation, especially odontogenic differentiation. We found that increased expression of JAB1 promoted odontogenic differentiation of DPSCs via Wnt/β-catenin signaling. The role of JAB1 in the odontogenic differentiation of DPSCs was further confirmed by knocking down JAB1. Our findings provide novel insights on odontogenic differentiation of DPSCs.

AGS3 is involved in TNF-α medicated osteogenic differentiation of human dental pulp stem cells.

Dental pulp stem cells (DPSCs) are multipotent adult stem cells capable of differentiating along the osteoblast, adipocyte, and chondrocyte lineages. Regulating differentiation of DPSCs may be a useful tool for regenerative medicine and cell-based therapy in oral diseases. Multisignaling pathways are involved in osteogenic differentiation of DPSCs. Recent studies show that cAMP/PKA/CREB signaling could stimulate the expression of genes such as bone morphogenic proteins 2 (BMP2), inhibitor of DNA binding 2 (ID2), bone sialoprotein, osteocalcin, and type XXIV collagen, which have been implicated in osteogenesis and bone formation. Activator of G-protein signaling 3 (AGS3, gene name G-protein signaling modulator-1, Gpsm1), an accessory protein for G-protein signaling, plays an important role in regulating the phosphorylation of cyclic AMP response element-binding protein (p-CREB). However, the involvement of AGS3 in osteogenic differentiation of DPSCs has not been explored. Our data indicated that increased expression of AGS3 would inhibit osteogenic differentiation of DPSCs exposed to inflammatory cytokine tumor necrosis factor α (TNF-α) via cAMP/PKA/CREB signaling. The negative role of AGS3 in osteogenic differentiation was further confirmed by knocking down and over expression of AGS3. Our findings may provide clinical implications for osteoporosis.