OBJECTIVETo isolate and cultivate human periodontal ligament stem cells (PDLSC) and to investigate the feasibility of PDLSC in vitro differentiation into chondrogenic phenotype.

METHODSPeriodontal tissue was obtained from healthy young human teeth extracted for orthodontic purposes. PDLSCs were isolated by single-colony selection and cultivated. PDLSC of passage 3 was plated at density of 1 x 10(7) cells/cm3 and induced with chondrogenic induction medium of DMEM containing TGF-beta1 (10 microg/L), IGF-1 (50 microg/L), dexamethasone (40 microg/L) and 10% FBS. In control group, the constructs were maintained in DMEM medium + 10% FBS. After 21 days induction, the results were evaluated by histology, histochemistry, immunohistochemistry and RT-PCR.

RESULTSThe constructs in experimental group were smooth and relatively firm in texture after 3 weeks of culture. Toluidine blue staining showed the formation of distinct lacuna structure. Positive staining of type II collagen was also detected by immunohistochemistry and it was confirmed by RT-PCR. In contrast, in the control group, the constructs collapsed gradually, lacuna was barely detected in histology and type II collagen expression negative.

CONCLUSIONSPeriodontal ligament contain stem cells can be isolated and cultivated. PDLSC have the potential of chondrogenic differentiation.

Adolescent ; Adult Stem Cells ; cytology ; Cell Differentiation ; Cells, Cultured ; Child ; Chondrocytes ; cytology ; Humans ; Periodontal Ligament ; cytology

OBJECTIVETo investigate the effects of 17beta-estradiol on the expression of alkaline phosphatase (ALP) and osteoprotegerin in human periodontal ligament cells.

METHODSHuman periodontal ligament cells (hPDLC) were obtained from periodontal tissue explants of teeth extracted for orthodontic treatment ALP activity was determined by PNPP, and OPG protein and corresponding mRNA levels were quantitatively detected by ELISA and RT-PCR RESULTS: ALP activity was significantly increased at 14 days and 21 days (P <0.05). 17beta-E2 of physiological concentration promoted secretion of OPG protein and expression of OPG mRNA (P <0.05). 17beta-E2 with high-dose showed no effect on OPG protein secretion and decrease OPG mRNA expression.

CONCLUSIONS17beta-E2 may have a positive impact on periodontium through promoting expression of ALP and OPG in hPDLC.

Alkaline Phosphatase ; metabolism ; Cells, Cultured ; Estradiol ; pharmacology ; Humans ; Osteoprotegerin ; metabolism ; Periodontal Ligament ; cytology ; drug effects ; metabolism

This study aims at exploring the effects of tensile strain and loading time on the secretion of IL-1 beta of human periodontal ligament fibroblast. Five tensile strain values including 0%, 8%, 12%, 16%, 20% and three loading time including 24 h, 48 h, 72 h are set in this study. The prepared cell samples are mounted on the self-devised loading apparatus in vitro. The content of IL-1 beta in each sample was determined using double-antibody ELISA. The secretion amount of IL-1 beta per day is directly proportional to the loading time and tensile strain value in tensile strain group of 8%, 12%, 16%. The secretion amount of IL-1 beta reaches its maximum at tensile strain value of 16% in loading time groups. Loaded strain for 24 h and 48 h, the secretion amount of IL-1 beta at tensile strain value of 20% is obviously more than that at value of 0%, but the amount already begin to decrease apparently. Loaded strain for 72 h, the secretion amount of IL-1 beta decreases to a great extent that it is less than the amount at strain value of 0%. The tensile strain stimulates the secretion of IL-1 beta by human periodontal ligament fibroblast when the strain is under normal physiological extent, but the stimulation effect fades out as time goes on.
Cells, Cultured ; Fibroblasts ; secretion ; Humans ; Interleukin-1 ; secretion ; Periodontal Ligament ; cytology ; secretion ; Tensile Strength ; Time Factors

Magnetic attachments have flux leakages, thus they will exert certain magnetic fields on the adjacent tissues when used in the patients' oral cavities. There are few research reports on the biological effects of the magnetic fields generated by magnetic attachments on human body. A cellular static magnetic field (SMF) loading system was developed in this study. By using this system, in vitro cultured human periodontal ligament cells (HPDLCs) were loaded with SMF simulating those of magnetic attachments. The cellular SMF loading system could produce constant SMF and the strength of the SMF is adjustable. The system is small and is able to exert SMF to cells cultured in different culture vessels such as culture dishes and culture plates, thus is suitable to researches in multiple biological items of cells. The results of the SMF loading experiment on HPDLCs showed that this cellular SMF loading system could effectively load cells with SMF of different strengths for different time in vitro. The development of this system has provided a useful tool for the researches on the cellular hiologioal effects of SMF.
Cells, Cultured ; Electromagnetic Fields ; adverse effects ; Humans ; Magnetics ; Periodontal Ligament ; cytology ; metabolism ; radiation effects

OBJECTIVETo explore the biological effects of tetracycline on cultured human periodontal ligament fibroblasts (HPDLFs).

METHODSIncreasing concentrations of tetracycline (1, 5, 20, 100, 500, 2500 microg/ml) were added to the medium of cultured HPDLFs, respectively. After co-incubated for 2 days, cell morphology was observed under reverse microscope, meanwhile, cell proliferation activity was assayed using MTT, the total amount of protein was detected with Coumassie Bright Blue method and DNA synthesis was measured by 3H-TdR.

RESULTSOver a concentration range of 1 to 100 microg/ml, cells demonstrated a normal appearance, spindle or fusiform shaped. Moreover, at a concentration range of 20 to 100 microg/ml, tetracycline significantly enhanced the proliferating activity and biosynthesis of HPDLFs (P < 0.01). However, higher concentration (2500 microg/ml) not only changed cell morphology, but also significantly inhibited cellular activity.

CONCLUSIONThe results suggested that proper doses of tetracycline could promote proliferation and biosynthesis of HPDLFs while higher concentrations of tetracycline had cytotoxic effect.

Cells, Cultured ; Fibroblasts ; drug effects ; Humans ; Periodontal Ligament ; cytology ; drug effects ; Tetracycline ; pharmacology

UNLABELLEDThe periodontal ligament fibroblast (PDLF) was cultivated artificially as the cell to be tested, and then it was loaded with mechanical stress-strain of different values and for different times. The cell and nucleus projected areas and shapes as well as the structure of cytoskeleton were tested by use of confocal laser scanning microscope and immunity fluorescence technique. Then the relationship among the stress-strain, the time, the shape and the structure of cytoskeleton of the PDLF was detected.

RESULTSIn the trial groups of 0, 8%, 12%, 16% strain values, the cell and nucleus projected areas were proportional to strain (stress) and time. The diameter, density and order of the structure of cytoskeleton increased in the strain and time dependent fashion. In the trial group of 20% strain values, the cell and nucleus projected areas decreased with the increase of time, and the structure of cytoskeleton became disorderly. It was demonstrated in this study that the shape and structure of cytoskeleton of PDLF underwent regular changes when the PDLF was loaded with the mechanical stress-strain.

Cells, Cultured ; Cytoskeleton ; physiology ; Fibroblasts ; cytology ; physiology ; Humans ; Periodontal Ligament ; cytology ; Radioimmunoassay ; Stress, Mechanical ; Tensile Strength ; physiology

OBJECTIVETo observe the effects of platelet-rich plasma (PRP) on inducing mineralization of human gingival fibroblasts (GF), periodontal ligament cells (PDLC) and alveolar bone osteoblasts (AOB).

METHODSHuman whole blood from healthy subjects was collected and PRP was obtained after twice centrifuged. GF, PDLC and AOB established from tissue explants were used at 4th passage in culture. All cells were seeded at density of 4 x 10(7)/L at 6 well tissue culture plate and cultured at standard condition for 96 hours, and then the different media was used. In the experimental group, the cells continued to be cultured in 50 ml/L PRP in DMEM media with mineral solution (10 mmol/L beta-glycerophosphate, 0.2 mmol/L freshly prepared ascorbic acid, 10 nmol/L dexamethasone), in the control group, the cells to be cultured in DMEM media with mineral solution, and in the blank group DMEM media was used. The media was changed every other day until day 30, the cells were dyed with von Kossa. Pictures were taken and analyzed by the image analysis software to semiquantitate the proportion of the positive area of von Kossa dye and attenuation aera of AOB.

RESULTSIn PDLC and AOB, the quantity of mineralization nodule in the experimental group and the control group was more than that in the blank group (P < 0.05), and there were more mineralization nodules in the experimental group than in the control group (P < 0.05), while the amount of mineralization nodules in GF was low in the three groups (P > 0.05). Attenuation of AOB in the experimental group was less than the control group (P < 0.05).

CONCLUSIONSPRP can enhance the quantity of mineralization in PDLC and AOB in vitro and reduce attenuation in AOB, which suggests that PRP may be helpful to periodontal alveolar bone mineralization and regeneration.

Adolescent ; Adult ; Alveolar Process ; cytology ; Cells, Cultured ; Female ; Fibroblasts ; chemistry ; cytology ; Gingiva ; cytology ; Humans ; Male ; Osteoblasts ; chemistry ; cytology ; Periodontal Ligament ; cytology ; Platelet-Rich Plasma ; Young Adult

Animals ; Bone Regeneration ; physiology ; Cell Differentiation ; Dental Enamel Proteins ; pharmacology ; Dental Pulp ; cytology ; Fibroblasts ; cytology ; Gingiva ; cytology ; Guided Tissue Regeneration, Periodontal ; methods ; Humans ; Induced Pluripotent Stem Cells ; cytology ; physiology ; Mouth Mucosa ; cytology ; Periodontal Ligament ; cytology ; Tissue Engineering ; methods

OBJECTIVETo examine whether human periodontal ligament fibroblasts (hPDLF) express neural growth factor (NGF).

METHODSPC12 cells were cultured in hPDLF-conditioned medium to evaluate the secretion of NGF by hPDLF.

RESULTShPDLF-conditioned medium resulted in neuronal differentiation and neurite outgrowth from PC12 cells. Only a small percentage of the cells grown in anti-NGF antiserum and DMEM expressed neuritis.

CONCLUSIONhPDLF are capable of synthesizing NGF and secreting it into the medium as the signaling molecules for innervation during the healing of the periodontal tissue.

Animals ; Cell Differentiation ; drug effects ; Cells, Cultured ; Culture Media, Conditioned ; pharmacology ; Fibroblasts ; cytology ; Humans ; Neurons ; cytology ; PC12 Cells ; Periodontal Ligament ; cytology ; Periodontium ; cytology ; Pheochromocytoma ; pathology ; Rats

OBJECTIVETo observe the effects of cells and scaffolds tissue engineering on the periodontal regeneration, and to evaluate the feasibility of nano-Hap-collagen (nHAC) as the scaffold material for periodontal tissue engineering.

METHODSDog autogenous periodontal ligament cells (PDLCs) cultured in vitro were collected and seeded on the three-dimensional framework of nHAC. The cell growth in the scaffolds was observed by scanning electron microscope. And then the PDLCs-nHAC composites were transplanted into man-made periodontal defects, and the groups filled with nothing or filled only with nHAC were the controls. The dogs were sacrificed after 8 weeks and the periodontal regeneration was observed histologically.

RESULTSScanning electron microscope showed the porous structure of nHAC and the eugonic growth of cells in the nHAC scaffolds. The histological observation showed that the PDLCs-nHAC groups exhibited more new bone, new periodontal ligament and new cementum occupying the majority of the defects than the control groups, and the epitheliums were not observed.

CONCLUSIONSPeriodontal regeneration could be enhanced by the cells and scaffolds tissue engineering, and the PDLCs and nHAC could be used as the seed cell and the scaffold material for periodontal tissue engineering.

Animals ; Biocompatible Materials ; Cells, Cultured ; Dogs ; Guided Tissue Regeneration ; Male ; Periodontal Attachment Loss ; surgery ; Periodontal Ligament ; cytology ; Regeneration ; Replantation ; methods ; Tissue Engineering ; methods ; Transfection