Tube Formation Capability and Chemotaxis of Skin Pericytes.

BACKGROUND: Pericytes (PCs), the critical components of vessels, are implicated in wound repair. This study aimed to explore the roles of PCs in wound healing and angiogenesis. METHODS: Skin PCs and human dermal microvascular endothelial cells (HDMECs) were isolated from patients' upper eyelid skin. Immunofluorescence staining was used to characterize the morphology of PCs. Tube formation and transwell chemotaxis assays were performed to explore PC's tube-forming capability and chemotaxis. Finally, we investigated the effects of PCs and endothelial cells on wound repair using skin wound of a rat model. RESULTS: Skin PCs exhibited a double-protrusion structure and characteristic antigen expression of neural/glial antigen 2 (NG2)+/platelet-derived growth factor receptor-ß (PDGFR-ß)+/alpha-smooth muscle actin (α-SMA)+/CD31-. Skin PCs could directly form lumen-like structures in a two dimensional (2D) culture environment, and mild hypoxia and starvation promoted the lumen-like structure formation. Furthermore, skin PCs quickly formed more stable lumen-like structures than HDMECs in matrigel, and they recruited HDMECs in a three dimensional (3D) culture environment. Transwell chemotaxis assay showed that PCs and HDMECs were chemotactic to each other. PCs could develop lumen-like structures in the skin wounds of rat models. The number of PCs mounted in wounded skin was compared to normal skin. The ratio of PCs to endothelial cells gradually increased after skin injury and reached its maximum on the 3rd day. CONCLUSIONS: Skin PCs have an excellent tube-forming capability and chemotaxis to endothelial cells. PCs might promote wound repair by recruiting endothelial cells.

Three New Niccolites: High-Temperature Phase Transitions, Prominent Anisotropic Thermal Expansions, Dielectric Anomalies, and Magnetism.

Three new iso-structural ammonium metal formates of [dmpnH2 ][M2 (HCOO)6 ], in which dmpnH2 2+ =N,N'-dimethyl-1,3-propylenediammoium and M=divalent Co, Zn and Mg ions, are reported. They possess niccolite metal formate frameworks with long-shaped cavities for the accommodation of dmpnH2 2+ cations. The three materials display reversible phase transitions of similar mechanism from ordered, antipolar or antiferroelectric, low-temperature phases in space group C2/c, to disordered, paraelectric, high-temperature phases in space group P 3 ‾ 1c, with quite high critical temperatures of 366, 370, and 334 K for Co, Zn, and Mg members, respectively. On warming, the dmpnH2 2+ cation experiences an ordered state with gradual increase of the local vibration motions of the central CH2 and terminal CH3 groups, a partially disordered state with gradually enhanced flipping motion between the major and minor orientations, and finally a twisting or rotating motion after the phase transition, accompanied by prominent anisotropic thermal expansions and dielectric anomalies/relaxations. The phase transition characters and relevant properties also exhibit a subtle metal-dependence. The Co member shows spin-canted antiferromagnetism below the Néel temperature of 16.1 K, with unusual large spontaneous magnetization and coercive field.

Psychological Stress Alters Extracellular Matrix Metabolism in Mandibular Condylar Cartilage.

OBJECTIVE: To explore the effect of long-term stress on the temporomandibular joint (TMJ) condyle and its possible underlying mechanism. METHODS: A 12-week, chronic unpredictable mild stress (CUMS) model was used to induce long-term psychological stress in rats. Rats were randomly divided into control group (CONT), chronic unpredictable mild stress group (CUMS) and chronic unpredictable mild stress with fluoxetine treatment group (CUMS + DT) (n = 30 per group). A 5 mg/kg dose of fluoxetine was intraperitoneally injected daily 0.5 h before stress. A sucrose preference test, plasma corticosterone test and open-field test were performed to verify the feasibility of the CUMS model. Histopathology was used to observe the pathological changes of condyle. The expression levels of inflammatory cytokines, matrix metalloproteases (MMPs) and extracellular matrix (ECM) were measured by real-time polymerase chain reaction, western blotting and immunohistochemistry. RESULTS: At 8 and 12 weeks after exposure to CUMS, the rats showed higher plasma corticosterone than the control rats. Additionally, for the open-field test, the rats exposed to CUMS spent more time in the centre zone and moved a shorter distance than the control and drug treatment rats. In addition, pathological changes in the condylar cartilage occurred in the 8-week CUMS subgroup and were more obvious in the 12-week CUMS subgroup. The CUMS caused an increase in the secretion of inflammatory cytokines, imbalanced expression of MMPs and tissue inhibitor of metalloproteinase-1 and accelerated degradation of ECM in condylar cartilage in a time-dependent manner. CONCLUSION: Osteoarthritis-like lesions can be caused by long-term CUMS in the mandibular condyles, which suggests that the imbalance in chondrocyte-secreted regulatory factors within the cartilage of the TMJ may play an important role in cartilage injury induced by psychological stress.

The Distinct Effects of Estrogen and Hydrostatic Pressure on Mesenchymal Stem Cells Differentiation: Involvement of Estrogen Receptor Signaling.

This study aimed to investigate the differential and synergistic effects of mechanical stimulation and estrogen on the proliferation and osteogenic or chondrogenic differentiation potential of bone marrow mesenchymal stem cells (BMSCs) and the roles of estrogen receptor (ER) in them. BMSCs were isolated and cultured using the whole bone marrow adherence method, and flow cytometry was used to identify the surface marker molecules of BMSCs. Cells were pre-treated with 1 nM 17ß-estradiol or 1 nM of the estrogen receptor antagonist tamoxifen. Then, the cells were stimulated with hydrostatic pressure. Assessment included flow cytometry analysis of the cell cycle; immunofluorescent staining for F-actin; protein quantification for MAPK protein; and mRNA analysis for Col I, OCN, OPN and BSP after osteogenic induction and Sox-9, Aggrecan and Col-II after chondrogenic induction. Hydrostatic pressure (90 kPa/1 h) and 1 nM 17ß-estradiol enhanced the cellular proliferation ability and the cytoskeleton activity but without synergistic biological effects. Estrogen activated ERKs and JNKs simultaneously and promoted the osteogenic differentiation, whereas the pressure just caused JNK-1/2 activation and promoted the chondrogenic differentiation of BMSCs. Estrogen had antagonism effect on chondrogenic promotion of hydrostatic pressure. Mechanobiological effects of hydrostatic pressure are closely associated with ERα activity. MAPK molecules and F-actin were likely to be important mediator molecules in the ER-mediated mechanotransduction of BMSCs.

Activation of satellite glial cells in the trigeminal ganglion contributes to masseter mechanical allodynia induced by restraint stress in rats.

It is commonly accepted that psychological stress contributes to the development of chronic orofacial pain. However, the neural mechanism underlying this process has remained unclear. The present study was performed to determine the involvement of satellite glia cells (SGCs) in the trigeminal ganglion (TG) in stress-induced increases in masseter muscle allodynia in rats. Using a chronic restraint stress model, we found that exposure to a 14-day stress but not a 3-day stress (6 h/day) caused decreased body weight gain, behavioral changes and marked masseter allodynia in rats. SGCs were dramatically activated, and substance P (SP) expression was significantly increased in the TG. A further analysis was undertaken to investigate the contribution of SGCs; the expression of interleukin-1ß (IL-1ß) in SGCs and interleukin-1 receptor I (IL-1RI) in neurons was significantly increased after chronic restraint stress, whereas injection of L-α-aminoadipate (a SGC inhibitor, LAA) into the TG dramatically inhibited the overexpression of these proteins. In addition, LAA or interleukin-1 receptor antagonist (IL-1ra) administration into the TG could significantly attenuate the mechanical masseter allodynia and overexpression of SP in the TG induced by restraint stress. These results suggest that SGC activation in the TG may play a role in masseter allodynia induced by restraint stress. The over-release of IL-1ß and excessive IL1-RI expressions have close relationship with the stress induced masseter allodynia.

Hydrostatic pressure promotes the proliferation and osteogenic/chondrogenic differentiation of mesenchymal stem cells: The roles of RhoA and Rac1.

Our previous studies have shown that hydrostatic pressure can serve as an active regulator for bone marrow mesenchymal stem cells (BMSCs). The current work further investigates the roles of cytoskeletal regulatory proteins Ras homolog gene family member A (RhoA) and Ras-related C3 botulinum toxin substrate 1 (Rac1) in hydrostatic pressure-related effects on BMSCs. Flow cytometry assays showed that the hydrostatic pressure promoted cell cycle initiation in a RhoA- and Rac1-dependent manner. Furthermore, fluorescence assays confirmed that RhoA played a positive and Rac1 displayed a negative role in the hydrostatic pressure-induced F-actin stress fiber assembly. Western blots suggested that RhoA and Rac1 play central roles in the pressure-inhibited ERK phosphorylation, and Rac1 but not RhoA was involved in the pressure-promoted JNK phosphorylation. Finally, real-time polymerase chain reaction (PCR) experiments showed that pressure promoted the expression of osteogenic marker genes in BMSCs at an early stage of osteogenic differentiation through the up-regulation of RhoA activity. Additionally, the PCR results showed that pressure enhanced the expression of chondrogenic marker genes in BMSCs during chondrogenic differentiation via the up-regulation of Rac1 activity. Collectively, our results suggested that RhoA and Rac1 are critical to the pressure-induced proliferation and differentiation, the stress fiber assembly, and MAPK activation in BMSCs.

Potential dental pulp revascularization and odonto-/osteogenic capacity of a novel transplant combined with dental pulp stem cells and platelet-rich fibrin.

Our aim is to investigate the cytobiological effects of autologous platelet-rich fibrin (PRF) on dental pulp stem cells (DPSCs) and to explore the ectopic and orthotopic possibilities of dental pulp revascularization and pulp-dentin complex regeneration along the root canal cavities of the tooth by using a novel tissue-engineered transplant composed of cell-sheet fragments of DPSCs and PRF granules. Canine DPSCs were isolated and characterized by assaying their colony-forming ability and by determining their cell surface markers and osteogenic/adipogenic differentiation potential. The biological effects of autologous PRF on DPSCs, including cell proliferation, alkaline phosphatase (ALP) activity and odonto-/osteogenic gene expression, were then investigated and quantified. A novel transplant consisting of cell-sheet fragments of DPSCs and PRF granules was adopted to regenerate pulp-dentin-like tissues in the root canal, both subcutaneously in nude mice and in the roots of canines. PRF promoted the proliferation of DPSCs in a dose- and time-dependent manner and induced the differentiation of DPSCs to odonto-/osteoblastic fates by increasing the expression of the Alp, Dspp, Dmp1 and Bsp genes. Transplantation of the DPSC/PRF construct led both to a favorable regeneration of homogeneous and compact pulp-like tissues with abundantly distributed blood capillaries and to the deposition of regenerated dentin along the intracanal walls at 8 weeks post-operation. Thus, the application of DPSC/PRF tissue constructs might serve as a potential therapy in regenerative endodontics for pulp revitalization or revascularization.

Involvement of trigeminal astrocyte activation in masseter hyperalgesia under stress.

It is commonly accepted that psychological stress contributes to the development of temporomandibular joint disorders, in which chronic orofacial pain is the main symptom. However, the central mechanism underlying the development of these disorders has remained unclear. The current study was performed to determine the involvement of the glia in the trigeminal spinal subnucleus caudalis in stress-induced increases in masseter muscle hyperalgesia in rats. After being subjected to chronic restraint stress, the animals showed decreased body weight gain, behavioral changes and marked masseter allodynia. We also found that astrocytes, but not microglia, in the trigeminal subnucleus caudalis (Vc) were dramatically activated. A further analysis was undertaken to investigate the contribution of the glia; we intrathecally injected l-α-aminoadipate (astrocyte-specific inhibitor) and/or minocycline (microglia-specific inhibitor) into the stressed rats. Our results showed that l-α-aminoadipate (LAA), but not minocycline, could significantly attenuate the mechanical masseter allodynia and behavioral changes induced by restraint stress. In addition, the expression of interleukin-1ß (IL-1ß) and phosphorylated N-methyl-d-aspartic acid receptor 1 (p-NR1) in the Vc was significantly increased after chronic restraint stress, whereas LAA dramatically inhibited the overexpression of IL-1ß and p-NR1. Taken together, these results suggest that activated astrocytes in the Vc may be one of the most important factors in the pathophysiology of masseter hyperalgesia induced by restraint stress and the following overexpression of IL-1ß and excessive NMDAR phosphorylation may ultimately contribute to masseter hyperalgesia. Thus, inhibiting spinal astrocytic activation may represent a novel therapeutic strategy for the treatment of orofacial pain induced by stress.

The combined use of cell sheet fragments of periodontal ligament stem cells and platelet-rich fibrin granules for avulsed tooth reimplantation.

The aim of this study was to construct a cell transplant method consisting of cell sheet fragments of periodontal ligament stem cells (PDLSCs) and platelet-rich fibrin (PRF) granules to enhance periodontal healing in avulsed tooth reimplantation. To test this concept in vitro, human PDLSCs were isolated and characterized by colony forming unit assay, cell surface marker characterizations, and their osteogenic/adipogenic differentiation potential. The biological effects of autologous PRF as a growth factor-enriched endogenous scaffold on human PDLSCs were then investigated and quantified for statistical analyses, including cell viability and proliferation, alkaline phosphatase (ALP) activity, and the gene expression of bone sialoprotein (BSP), osteocalcin (OCN), collagen I (Col-I), and cementum protein 23 (CP23). It was found that the PRF induced a significant and continuous stimulation of proliferation in human PDLSCs throughout the 7-day incubation period. Furthermore, the PRF suppressed the osteoblastic differentiation of PDLSCs by decreasing ALP activity and the gene expression of BSP and OCN while up-regulating the mRNA expression levels of Col-I and CP23 during the testing period. To assess the potential application of the PDLSCs/PRF construct in tooth reimplantation, 36 incisors were extracted from 6 dogs. The incisors then underwent 2 h of dry storage and were randomly divided into four groups receiving different strategies of reimplantation, where the avulsed teeth were reimplanted with the use of the autologous PDLSCs/PRF construct (cell sheet fragments in combination with PRF granules), with the use of autologous PDLSCs or PRF alone, or without adjuvant use of PRF or PDLSCs. Eight weeks post-reimplantation, the PDLSCs/PRF group achieved a more effective periodontal healing, characterized by the regeneration of PDL-like tissues and a reduction of ankylosis and inflammation, compared with the other testing groups. These overall results suggest that the PDLSCs/PRF construct may be a useful tool for alveolar surgery that has the potential to improve the clinical outcomes in future avulsed tooth reimplantations.

6-Nitro-1,3-benzothia-zole-2(3H)-thione.

In the title mol-ecule, C(7)H(4)N(2)O(2)S(2), the nitro group is twisted by 5.5 (1)° from the plane of the attached benzene ring. In the crystal, N-H⋯S hydrogen bonds link pairs of mol-ecules into inversion dimers, which are linked by weak C-H⋯O inter-actions into sheets parallel to (101). The crystal packing exhibits short inter-molecular S⋯O contacts of 3.054 (4) Šand π-π inter-actions of 3.588 (5) Šbetween the centroids of the five- and six-membered rings of neighbouring mol-ecules.