OBJECTIVETo compared the difference in contact guidance activity among microgroove surfaces with different sizes of human gingival fibroblast (HGF), with the hope of providing basis for size selection of microgroove for transmucosal part of dental implant.

METHODSBasing on the size of HGF, microgroove titanium surfaces were fabricated by photolithography with parallel grooves: 15, 30 or 60 µm in width and 5 or 10 µm in depth. The groups that used different microgroove surfaces were denoted as T15/5, T15/10, T30/5, T30/10, T60/5, and T60/10. Group T0 (the control meanwhile was a sputter of titanium on a simple planar silicon substrate). The morphology that HGF arranged along the groove was analyzed by immunofluorescence staining. Difference in contact guidance activity was quantitatively compared basing on the consistency of nucleus arrangement and deformation ratio.

RESULTSMicrogroove groups had significantly higher consistency of nucleus arrangement and deformation ratio compared to the control group, with T60/10 had the highest consistency of 0.937±0.024, and T15/5 had the lowest consistency of 0.660±0.016 and T60/10 had the highest deformation ratio of 3.555±0.205, and T15/5 had the lowest deformation ratio of 1.819±0.011.

CONCLUSIONSMicrogroove surfaces of all the different sizes show contact guidance activity on HGF, and the contact guidance activity increases with the increase of width and depth.

Cell Adhesion ; physiology ; Dental Implants ; Fibroblasts ; cytology ; physiology ; Gingiva ; cytology ; Humans ; Surface Properties ; Titanium

Animals ; Endothelial Cells ; metabolism ; physiology ; Fibroblasts ; metabolism ; physiology ; Humans ; Hypertension, Pulmonary ; etiology ; genetics ; metabolism ; Myocytes, Smooth Muscle ; metabolism ; physiology

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

One of the differences between fetal and adult skin healing is the ability of fetal wounds heal without contraction and scar formation. Extracellular matrix (ECM) provides a substratum for cells adhesion, migration, and proliferation and can directly influence the form and function of cells. As motility is essential for many important biological events, including wound healing, inflammatory response, embryonic development, and tumor metastasis, this study was designed to compare the motilities cultured dermal fetal and neonatal fibroblasts in the extracellular matrix. The motility of cultured fetal and neonatal fibroblasts was compared using a video-microscopy system that was developed in combination with a self-designed CO2 mini-incubator. To determine migration speed, cells were viewed with a 4X phase-contrast lens and video recorded. Images were captured using a color CCD camera and saved in 8-bit full-color mode. We found that cultured fetal fibroblasts move faster than neonatal fibroblast on type I collagen (fetal fibroblast, 15.1 micrometer/hr; neonatal fibroblast, 13.7 micrometer/hr), and in fibronectin (fetal fibroblast, 13.2 micrometer/hr; neonatal fibroblast, 13.0 micrometer/hr) and hyaluronic acid (fetal fibroblast, 11 micrometer/hr; neonatal fibroblast, 9.8 micrometer/hr).
Cell Movement ; Cells, Cultured ; Comparative Study ; Extracellular Matrix/*physiology ; Fetus/physiology ; Fibroblasts/*physiology ; Human ; Infant, Newborn ; Skin/cytology/*embryology ; *Skin Physiology

UNLABELLEDObtaining human blastocysts is a prerequisite for cell replacement therapy using embryonic stem cells. We established an interspecies somatic cell nuclear transfer (iSCNT) technique for producing blastocysts without sacrificing human oocytes. Human foetal fibroblasts were used as donor cells injected into the enucleated bovine oocytes in nuclear transfer, whereas bovine foetal fibroblasts were used to produce intraspecies embryos. We also examined the fate of human and bovine mitochondrial DNA (mtDNA) during preimplantation development after nuclear transfer by PCR. PCR analysis for the detection of human and bovine mtDNA was done at the 2,8-morula, and blastocyst stages of the embryos.

RESULT2.8% interspecies embryos developed to blastocysts after cultured in an SOF medium, while blastocyst rate of intraspecies embryos were 10.1%. Both human and bovine mtDNAs existed until the morula stage, whereas only the bovine mtDNA was found at the blastocyst stage. These results indicated that interspecies cloning without using human oocytes could generate human blastocysts. Because of the incoordination between bovine mtDNA and human nuclear gene, developmental rate of interspecies embryos was significantly lower than intraspecie. Whether the embryonic stem cell could be used for cell replacement therapy need further research.

Animals ; Blastocyst ; cytology ; physiology ; Cattle ; Cloning, Organism ; DNA, Mitochondrial ; genetics ; Embryonic Development ; physiology ; Female ; Fibroblasts ; physiology ; Humans ; Nuclear Transfer Techniques ; Oocytes ; physiology ; Species Specificity

Objective: To investigate the regulatory effects of bio-intensity electric field on the transformation of human skin fibroblasts (HSFs). Methods: The experimental research methods were used. HSFs were collected and divided into 200 mV/mm electric field group treated with 200 mV/mm electric field for 6 h and simulated electric field group placed in the electric field device without electricity for 6 h. Changes in morphology and arrangement of cells were observed in the living cell workstation; the number of cells at 0 and 6 h of treatment was recorded, and the rate of change in cell number was calculated; the direction of cell movement, movement velocity, and trajectory velocity within 3 h were observed and calculated (the number of samples was 34 in the simulated electric field group and 30 in 200 mV/mm electric field group in the aforementioned experiments); the protein expression of α-smooth muscle actin (α-SMA) in cells after 3 h of treatment was detected by immunofluorescence method (the number of sample was 3). HSFs were collected and divided into simulated electric field group placed in the electric field device without electricity for 3 h, and 100 mV/mm electric field group, 200 mV/mm electric field group, and 400 mV/mm electric field group which were treated with electric fields of corresponding intensities for 3 h. Besides, HSFs were divided into simulated electric field group placed in the electric field device without electricity for 6 h, and electric field treatment 1 h group, electric field treatment 3 h group, and electric field treatment 6 h group treated with 200 mV/mm electric field for corresponding time. The protein expressions of α-SMA and proliferating cell nuclear antigen (PCNA) were detected by Western blotting (the number of sample was 3). Data were statistically analyzed with Mann-Whitney U test, one-way analysis of variance, independent sample t test, and least significant difference test. Results: After 6 h of treatment, compared with that in simulated electric field group, the cells in 200 mV/mm electric field group were elongated in shape and locally adhered; the cells in simulated electric field group were randomly arranged, while the cells in 200 mV/mm electric field group were arranged in a regular longitudinal direction; the change rates in the number of cells in the two groups were similar (P>0.05). Within 3 h of treatment, the cells in 200 mV/mm electric field group had an obvious tendency to move toward the positive electrode, and the cells in simulated electric field group moved around the origin; compared with those in simulated electric field group, the movement velocity and trajectory velocity of the cells in 200 mV/mm electric field group were increased significantly (with Z values of -5.33 and -5.41, respectively, P<0.01), and the directionality was significantly enhanced (Z=-4.39, P<0.01). After 3 h of treatment, the protein expression of α-SMA of cells in 200 mV/mm electric field group was significantly higher than that in simulated electric field group (t=-9.81, P<0.01). After 3 h of treatment, the protein expressions of α-SMA of cells in 100 mV/mm electric field group, 200 mV/mm electric field group, and 400 mV/mm electric field group were 1.195±0.057, 1.606±0.041, and 1.616±0.039, respectively, which were significantly more than 0.649±0.028 in simulated electric field group (P<0.01). Compared with that in 100 mV/mm electric field group, the protein expressions of α-SMA of cells in 200 mV/mm electric field group and 400 mV/mm electric field group were significantly increased (P<0.01). The protein expressions of α-SMA of cells in electric field treatment 1 h group, electric field treatment 3 h group, and electric field treatment 6 h group were 0.730±0.032, 1.561±0.031, and 1.553±0.045, respectively, significantly more than 0.464±0.020 in simulated electric field group (P<0.01). Compared with that in electric field treatment 1 h group, the protein expressions of α-SMA in electric field treatment 3 h group and electric field treatment 6 h group were significantly increased (P<0.01). After 3 h of treatment, compared with that in simulated electric field group, the protein expressions of PCNA of cells in 100 mV/mm electric field group, 200 mV/mm electric field group, and 400 mV/mm electric field group were significantly decreased (P<0.05 or P<0.01); compared with that in 100 mV/mm electric field group, the protein expressions of PCNA of cells in 200 mV/mm electric field group and 400 mV/mm electric field group were significantly decreased (P<0.05 or P<0.01); compared with that in 200 mV/mm electric field group, the protein expression of PCNA of cells in 400 mV/mm electric field group was significantly decreased (P<0.01). Compared with that in simulated electric field group, the protein expressions of PCNA of cells in electric field treatment 1 h group, electric field treatment 3 h group, and electric field treatment 6 h group were significantly decreased (P<0.01); compared with that in electric field treatment 1 h group, the protein expressions of PCNA of cells in electric field treatment 3 h group and electric field treatment 6 h group were significantly decreased (P<0.05 or P<0.01); compared with that in electric field treatment 3 h group, the protein expression of PCNA of cells in electric field treatment 6 h group was significantly decreased (P<0.01). Conclusions: The bio-intensity electric field can induce the migration of HSFs and promote the transformation of fibroblasts to myofibroblasts, and the transformation displays certain dependence on the time and intensity of electric field.
Actins/biosynthesis* ; Cell Differentiation/physiology* ; Cell Movement/physiology* ; Electric Stimulation Therapy ; Electricity ; Fibroblasts/physiology* ; Humans ; Myofibroblasts/physiology* ; Proliferating Cell Nuclear Antigen/biosynthesis* ; Skin/cytology*

AIMTo explore the effects of tRNA on the growth of mammalian cells.

METHODSL929, NIH3T3, MCF-7 and PC12 cells were seeded in 96 well culture plate individually, and incubated at 37 degrees C in 5% CO2 for 4 h, the tRNAs from different species were added to the culture media individually. After certain time of incubation, the viability of the cells was evaluated by the MTT methods. Sub-confluent L929 cells were incubated with 200 microg/ml ytRNA for different times, then the cells were pooled and analyzed with flow cytometry assay.

RESULTStRNA specifically inhibited the growth of L929 cells in a dose-dependent manner. The sizes of tRNA-treated cells showed larger sizes and longer processes than those of untreated cells. Flow cytometric analysis further showed that most of tRNA-treated cells were arrested in S phase of the cell cycle.

CONCLUSIONThe cell growth inhibitory effects of tRNAs were caused mainly by their degraded fragments. The results suggested that tRNA or its degraded fragments might play important roles in regulation of cell proliferation.

Animals ; Cell Cycle Checkpoints ; physiology ; Cell Line ; Cell Proliferation ; Fibroblasts ; cytology ; Flow Cytometry ; Mice ; RNA, Transfer ; physiology

OBJECTIVETo observe the effects of supernatants of normal skin keratinocytes(NK) and scar keratinocytes(SK) on proliferation and collagen synthesis of hypertrophic scar fibroblasts(HSFB).

METHODSThe supernatant, collected from cultured NK and SK, was added to the cultivated HSFB. The MTT-method, 3H-proline incorporation and radioimmunoassay were employed to measure the cell proliferation, collagen synthesis and secretion.

RESULTSNK supernatant could inhibit HSFB proliferation and increase the collagen synthesis, but inhibit collagen secretion, as compared with the control group. On the contrary, SK supernatant could increase collagen synthesis and secretion, which had little effects on HSFB proliferation.

CONCLUSIONKeratinocytes derived from normal skin and hypertrophic scar show different effects on hypertrophic scar fibroblasts.

Cell Division ; Cells, Cultured ; Cicatrix, Hypertrophic ; metabolism ; pathology ; Collagen ; biosynthesis ; Culture Media, Conditioned ; Fibroblasts ; physiology ; Humans ; Keratinocytes ; physiology

We describe three cases of morphea which present lesions of the disease at the site of mechanical traumas. In these patients, local traumatization of cutaneous tissue appear to be the initiating event for the developrnent of new lesions (isomorphic phenomenon) associated with their possible morphea prone constitutions. The mechanism involving these local stimuli responsible for the excessive collagen production is lifficult to explain. Upon biologic stress or trauma, normal cutaneous physiology may cause loca production and release of inflammatory mediators/cytokines, such as transforrning growth factor-beta(the major connective tissue growth factor-inducer). Perhaps, possible dysregulation of collagen-growth or fibrosis- promoting cytokines causing some clonal overactivity of the fibroblast, or with other undefined mechanisms may cause excessive synthesis of collagen to the local effects of such triggering factors in susceptible/subclinical individuals. Although no definite environrriental/physical influences on the developrnent of morphea have been described, rnechanical trauma niay occasionally be regarded as a contributing factor as seen in these patients.
Collagen ; Connective Tissue ; Constitution and Bylaws ; Cytokines ; Fibroblasts ; Humans ; Physiology ; Scleroderma, Localized*

A hyaluronic acid (HA) incorporated porous collagen matrix was fabricated at -70 degree C by lyophilization. The HA incorporated collagen matrix showed increased pore size in comparison with collagen matrix. Biodegradability and mechanical properties of matrices were controllable by varying the ultraviolet (UV) irradiation time for cross-linking collagen molecules. Addition of HA to collagen matrix did not effect ultimate tensile stress after UV irradiation. HA incorporated collagen matrices demonstrated a higher resistance against the collagenase degradation than collagen matrix. In an in vitro investigation of cellular behavior using dermal fibroblasts on the porous matrix, HA incorporated collagen matrix induced increased dermal fibroblast migration and proliferation in comparison with collagen matrix. These results suggest that the HA incorporated collagen porous matrix assumes to enhance dermal fibroblast adaptation and regenerative potential.
Collagen/*metabolism ; Extracellular Matrix/*metabolism ; Fibroblasts/*physiology ; Human ; Hyaluronic Acid/*metabolism ; Porosity