Evaluation of osteogenic activity of periosteal-derived cells treated with inflammatory cytokines

INTRODUCTION: Skeletal homeostasis is normally maintained by the stability between bone formation by osteoblasts and bone resorption by osteoclasts. However, the correlation between the inflammatory reaction and osteoblastic differentiation of cultured osteoprogenitor cells has not been fully investigated. This study examined the effects of inflammatory cytokines on the osteoblastic differentiation of cultured human periosteal-derived cells. MATERIALS AND METHODS: Periosteal-derived cells were obtained from the mandibular periosteum and introduced into the cell culture. After passage 3, the periosteal-derived cells were further cultured in an osteogenic induction Dulbecco's modified Eagle's medium (DMEM) medium containing dexamethasone, ascorbic acid, and beta-glycerophosphate. In this culture medium, tumor necrosis factor (TNF)-alpha with different concentrations (0.1, 1, and 10 ng/mL) or interleukin (IL)-1beta with different concentrations (0.01, 0.1, and 1 ng/mL) were added. RESULTS: Both TNF-alpha and IL-1beta stimulated alkaline phosphatase (ALP) expression in the periosteal-derived cells. TNF-alpha and IL-1beta increased the level of ALP expression in a dose-dependent manner. Both TNF-alpha and IL-1beta also increased the level of alizarin red S staining in a dose-dependent manner during osteoblastic differentiation of cultured human periosteal-derived cells. CONCLUSION: These results suggest that inflammatory cytokines TNF-alpha and IL-1beta can stimulate the osteoblastic activity of cultured human periosteal-derived cells.

Effects of Repeated Dosing of 6% Hydroxyethyl Starch or 7.5% Sodium Chloride on Inflammatory Response and Oxidative Damage in Experimental Intracerebral Hemorrhage Rats / 中国康复理论与实践

@#Objective To observe the effects of repeated dosing of 6% hydroxyethyl starch (130/0.4) or 7.5% sodium chloride on brain edema after experimental intracerebral hemorrhage (ICH) in rats. Methods 167 male SD rats were divided into four groups randomly: Sham operation group (S, n=20), ICH control group (M, n=38), 7.5% sodium chloride group (N, n=55) and 6% hydroxyethyl starch group (H, n=54). The model of the ICH was established with stereotactically infusing 50 μl of the autologous femoral artery blood into the right caudate nucleus. group N and group H received 7.5% sodium chloride 5 ml/kg and 6% hydroxyethyl starch 30 ml/kg at 2 h, 24 h, 48 h and 72 h after operation respectively. The tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), superoxide dismutase (SOD) and malondialdehyde (MDA) in the tissue around the hemorrhage were measured at different time point. Results The IL-6 in group N was significantly more than that in group M at 24 h and 72 h after infusion (P<0.05), and the TNF-α in group H was less than that in group M at 24 h and 48 h after infusion (P<0.05). The SOD in group M decreased to the bottom at 48 h and 72h after ICH. SOD in group N and group H at 24 h, 48 h and 72 h after infusion was both significant more than that in group M (P<0.05). MDA in group H at 72 h after infusion was less than that in group M (P<0.05). Conclusion Repeated infusion of 6% hydroxyethyl starch (130/0.4) or 7.5% sodium chloride can decrease inflammatory response of brain tissue after ICH, which may protect brain from oxidative damage.

Relationship between Tumor Necrosis Factor-alpha and Smoking-related Chronic Obstructive Pulmonary Disease / 中国康复理论与实践

@#Objective To investigate the relationship between tumor necrosis factor-alpha (TNF-α) and development of chronic obstructive pulmonary disease (COPD).Methods COPD patients and controls were divided into three groups: COPD group (n=66), smoker control group (n=42) and health control group (n=23). COPD group was further divided into the serious group (n=23) and non-serious group (n=43). The concentration of TNF-α of all cases was detected by human Th1/Th2 cytokine kit.Results The concentration of TNF-α in the COPD group was significantly higher than that of the smoker and healthy groups ( P<0.01). Furthermore, compared to non-serious COPD group, the concentration of TNF-α was higher in the serious COPD group ( P<0.05).Conclusion The concentration of TNF-α might be related with the pathogenesis and development of COPD.

Expression of angiogenin, TGF-beta, VEGF, APEX and TNF-alpha in oral squamous cell carcinoma

PURPOSE: The purpose of this study was to verify that the expressions of angiogenin, transforming growth factor-beta(TGF-beta), vascular endothelial growth factor(VEGF), human apurinic/apyrimidinic endonuclease(APEX) and tumor necrosis factoralpha(TNF-alpha) were associated with the tumorigenesis of the oral squamous cell carcinoma(OSCC). MATERIALS AND METHODS: Fifty-one samples of OSCC and fifteen normal oral mucosae were obtained to analyze the expression levels of above five factors. mRNA expressions were quantified by the quantitative competitive PCR(QC-PCR) method. After 2% agarose gel electrophoresis stained with ethidium bromide, the concentration of mRNA was calculated by a digital image analysis system. The expression levels of angiogenin, TGF-beta, VEGF, APEX and TNF-alpha were compared by unpaired Student's ttests between cancer and normal tissues. We analyzed statistically to find the cut-off values that would be useful as diagnostic markers, and the linear regression analysis between every two factors of these five factors by SAS system. RESULTS: All of these five factors (angiogenin: P<0.0037, TGF-beta: P<0.0001, VEGF: P<0.0102, APEX: P<0.0023, TNF-alpha: P<0.0074) were significantly correlated with OSCC. In the analysis to find the cut-off values for the diagnosis, we could not find any value that had a reasonable sensitivity and specificity. In the linear regression analysis, there were correlations between angiogenin and TNF-alpha, TGF-beta and VEGF, TGF-beta and APEX, TGF-beta and TNF-alpha, VEGF and APEX, VEGF and TNF-alpha, APEX and TNF-alpha. CONCLUSION: Our results suggest that not only angiogenin, TGF-beta, VEGF, APEX and TNF-alpha are significantly associated with the tumorigenesis, but also the close relationship between these factors might enhance the tumorigenesis of OSCC. We can not find clinical availability for diagnosis.

The Effects of Air-borne Particulate Matters on the Alveolar Macrophages for the TNF-alpha and IL-1beta Secretion / 결핵및호흡기질환

BACKGROUND: PM is known to induce various pulmonary diseases, including asthma, cancer, fibrosis and chronic bronchitis. Despite the epidemiological evidence the pathogenesis of PM-related pulmonary diseases is unclear. METHODS: This study examined the effects of PM exposure on the secretion of TNF-alpha and IL-1beta in the cultured alveolar macrophages. The cultured primary alveolar macrophages were treated with the medium, PM (5~20 microgram/cm2), LPS (5ng/ml), and PM with LPS for 24h and 48h respectively. ELISA was used to assay the secreted TNF-alpha and IL-beta in the culture medium. Western blotting was used to identify and determine the level of proteins isolated from the culture cells. The cells cultured in the Lab-Tek(R) chamber slides were stained with immunocytochemical stains. RESULTS: PM induced TNF-alpha and IL-1beta secretion in the culturing alveolar macrophages, collected from the SPF and inflammatory rats. However, the effects were only dose-dependent in the inflammatory macrophages. When the cells were co-treated with PM and LPS, there was a significant synergistic effect compared with the LPS in the both cell types. CONCLUSION: PM might be play an important role in the induction and/or potentiation of various lung diseases by oversecretion of TNF-alpha and IL-1beta.

TNF-alpha and IL-12 Secretion in Macrophages in Response to Spores of Bacillus anthracis Sterne

Baerobic, spore forming, and rod-shaped bacterium. Anthrax spores are introduced into macrophage by phagocytosis and multiply after germination. The anthrax spores infected in macrophage produce lethal toxin eventually caused cell death. In this study, we analyzed apoptosis and cytokine TNF-alpha and IL-12 secretion after the infection of spores of B. anthracis Sterne in the murine macrophage RAW264.7 cells and in the primary human macrophages. In murine macrophage RAW264.7 cells infected by spore of B. anthracis Sterne, the cells were markedly changed in secretion of TNF-alpha (482~6,213 pg/ml) by lethal toxin, and induced apoptosis. In case of RAW264.7 cells infected by formalin-inactivated spores of B. anthracis, the cells were not able to produce lethal toxin, which released lower level concentration of TNF-alpha (7.7~97.2 pg/ml), and rarely induced apoptosis. When primary human macrophage cells infected with spores of B. anthracis Sterne, they secreted TNF-alpha (5~16 pg/ml), and induced apoptosis about 1% of total cells. We presented that inducing apoptosis by spores of B. anthracis Sterne capable of expressing lethal toxin is related with the secretion of TNF-alpha in murine macrophage RAW264.7 cells. These studies revealed that human and murine macrophages has affected differently by anthrax lethal toxin produced by spores of B. anthracis Sterne.

The effect of neuropeptides on secretion of Interleukin-8 (IL-8) / 대한치과보존학회지

We investigated the secretion of Interleukin-8 (IL-8) from ginviva and periodontal ligament stimulated with Substance P (SP) and Calcitonin Gene-related Peptide (CGRP). Gingiva (GF), periodontal ligament (PDLF) and pulp (PF) tissues were collected from extracted intact 3rd molars. Cultured cells were stimulated with different concentrations of SP for 4 hrs, and stimulated with SP, CGRP and Tumor Necrosis Factor-alpha (TNF-alpha) for 8 hrs. Then RNase Protection Assay was carried out. ELISA was performed using supernatants of stimulated cells for quantitative analysis of IL-8. Results were assessed using student t-test with significance of P < 0.05. According to this study, the results were as follows: 1. IL-8 mRNA was detected in all type of cells studied (PF, GF and PDLF). 2. IL-8 mRNA expression was not increased after stimulating 4 hrs with SP (10(-5)M) and SP (10(-8)M) compared with Mock stimulation in all type of cells studied. 3. IL-8 mRNA expression was not increased after stimulating 8 hrs with SP (10(-4)M) and CGRP (10(-6)M) compared with Mock stimulation in all type of cells studied. 4. TNF-alpha(2 ng/ml) increased the expression of IL-8 mRNA in all kind of cells studied. 5. The secretion of IL-8 from GF was increased 8 hrs after the stimulation with CGRP (10(-6)M) (p < 0.05). 6. The secretion of IL-8 from PDLF was increased 8 hrs after the stimulation with SP (10(-4)M) (p < 0.05). Calcitonin Gene-related Peptide (CGRP) increased Interleukin-8 (IL-8) which plays an important role in chemotaxis of neutrophil in Calcitonin Gene-related Peptide (CGRP) gingival tissue, whereas Substance P increased the secretion of IL-8 from periodontal ligament.

Association between chronic hepatitis B virus infection and tumor necrosis factor-alpha gene promoter polymorphisms / 대한내과학회지

BACKGROUND: The reasons for the viral persistence and disease progression of hepatitis B virus (HBV) infection are unknown, but are probably related to host immune factors. Cytokines such as tumor necrosis factor-alpha (TNF-alpha) play significant roles in inflammatory and immune defense. This study was undertaken to investigate the association between HBV infection and polymorphisms of TNF-alpha gene promoter polymorphism. METHODS: We studied 412 Korean patients with chronic HBV infection (72 inactive carriers, 261 chronic hepatitis, 79 liver cirrhosis) and 204 healthy individuals who recovered from HBV infection. We assessed two biallelic polymorphisms in TNF-alpha gene promoter (at position -308, -238) by single base primer extension assay (SNP ITTM). RESULTS: Genotype frequencies of TNF-alpha gene promoter at position -308 and -238 were not different between the clearance and the persistence group in univariate analysis. In multivariate analysis after adjusting age and sex, TNF 308 G/G genotype was associated with HBV persistence (ORs;1.71, p=0.039). Moreover, concerning the haplotype analysis, -308G/ -238G homozygotes showed much higher correlation with HBV persistence (ORs;1.88, p=0.005). Genotype distributions of both gene promoters in inactive carriers were similar to those in patients with chronic progressive liver disease (chronic hepatitis and liver cirrhosis). CONCLUSION: The carriers of -308 G/G genotype and -308G / -238G haplotype homozygotes in the TNF-alpha promoter region have higher risk of persistent HBV infection.

Tumor Necrosis Factor- and Resorption of Calcium Sulfate Used as a Bone Graft Substitute in Spinal Fusion in Rabbits / 대한정형외과학회잡지

PURPOSE: To investigate the effects of tumor necrosis factor-alpha on calcium sulfate, as a bone graft substitute, in terms of achieving inter-transverse fusion in experimental rabbits. MATERIALS AND METHODS: Twenty adult New Zealand white rabbits were used in our study. 0.4 mg of calcium sulfate was mixed with 0.4 mg of autogenous iliac bone and grafted on the left intertransverse space of L4-5 or L5-6, and then 0.8 mg of autogenous iliac bone was grafted on the right side at the same level. Thus, the experimental rabbits served as their own control. At postoperative 0, 4, 8, 12, and 16 weeks, plain roentgenography was performed to evaluate the bony union. At 16 weeks, all rabbits were sacrificed and histologic evidences of the bony union observed by H and E and trichrome staining. Computed tomography was also used to evaluate the union state. An immuno-histochemical study for TNF-alpha was to investigate the union. RESULTS: Bone graft, mixed with calcium sulfate was resorbed completely radiologically and histologically in 20 rabbits (100%). In contrast, a graft using autogenous cancellous bone showed complete bony union in 15 of 20 rabbits (75%). By immuno-histochemical staining, TNF-alpha was detected at the calcium sulfate mixed autogenous bone graft site. CONCLUSION: Union was not showed at the bone graft site, mixed with calcium sulfate. Therefore, TNF-alpha plays an important role causing subsequent calcium resorption.

Immunohistochemical studies of Interleukin-8 and Tumor necrosis factor-alpha on psoriatic lesions / 대한피부과학회지

BACKGROUND: Psoriasis is a chronic skin disease characterized histologically by prominent keratinocyte hyperplasia and an early inflammatory cell infiltrate. However, the pathogenesis is not fully understood yet. Recently, there has been growing interest in the probable role of a T cell mediated immune response in the pathogenesis. Especially, cytokine network involved in psoriasis has been studied. OBJECTIVE: This study was done to investigate the role of Interleukin-8(IL-8) and its inducer, Tumor necrosis factor-alpha(TNF-alpha) for pathogenesis of psoriasis. METHODS: We performed immunohistochemical staining using antibodies for IL-8 and TNF-alpha in frozen skin samples of 14 psoriatic patients who had not been treated for psoriatic lesion for 1 month and 3 normal skin samples as controls. RESULTS: There was no detectable discrete immunoreactivity for either TNF-alpha and IL-8 in epidermis and dermis of normal controls. In psoriatic lesions, the expression of TNF-alpha was found in dermal cells within the papillary dermis, particularly around blood vessels. The expression of IL-8 was presented on suprabasal keratinocytes above TNF-alpha-positive dermal tissue. CONCLUSION: Our results strongly suggest that histologic expression and functional interaction between IL-8 and its inducer, TNF-alpha have significant roles in the pathogenesis of psoriasis.

Effect of Retinoic Acid on Growth and Transduced Tumor Necrosis Factor-alpha Gene Expression of Human Bladder Tumor Cell Lines / 대한비뇨기과학회지

INTRODUCTION AND OBJECTIVES: Retinoic acid (RA) is known as a potent chemopreventive agent in bladder tumor. Recently, RA has gained attention for up-regulation of transduced gene expression via long terminal repeat (LTR) transcriptional promotion. In this study, we investigated the possible dual effect of RA, growth inhibition and up-regulation of transduced gene expression which contains LTR promoter in human bladder carcinoma cell lines. MATERIALS AND METHODS: Human bladder carcinoma cell lines CY-24, J-82, HT-1197, ATCC) were transduced with Moloney murine leukemia virus containing cDNA of TNF-alpha. The growth of transduced and parent cell line was measured by tetrazolium based colorimetric assay (MTF). Transduced TNF-alpha gene expression was determined by ELISA method. RESULTS: TNF-alpha production was increased approximately twofold after treatment with RA (10 uM) in all three cell lines. This increase was dependent on RA concentration. RA treatment of transduced and parent cell line resulted in dose dependent inhibition of cell proliferation(up to 80% inhibitionwith 10 uM RA) in all parental and transduced cell lines. CONCLUSIONS: These results indicate that RA shows dual effect in cytokine gene transduced bladder carcinoma cells with retroviral vector containing LTR promoter and could be a supplement to the gene therapy of bladder cancer.

Effect of Transduced Tumor Necrosis Factor-alpha Gene Expression on Growth of Human Bladder Tumor Cell Lines / 대한비뇨기과학회지

OBJECTIVE: To evaluate the effect of transduced tumor necrosis factor-a(TNF-a) gene expression on growth of human bladder tumor cell lines in vitro. MATERIALS AND METHODS: The complete cDNA of TNF-a was introduced to three human bladder tumor cell lines(F-24, J-82, HT-1197) using a retroviral vector, a recombinant form of Molony murine leukemia virus with TNF-a and Neo gene and transfected cells were selected by exposure to neomycin analog G418. Gene transfer and expression were confirmed by polymerase chain reaction(PCR) and reverse transcription polymerase chain reaction(RT-PCR)-Southern blotting. Cell growth was measured by MTT assay Result is Successful gene transfer and expression were confirmed in all three cell bladder tumor lines. Growth of transfected cells were compared with parental cell lines and no differences were found in all three cell lines(p>0.05). CONCLUSION: Expression of transduced TNF-t gene could not show any effect on growth of human bladder tumor cells in vitro.

Effects suramin, tumor necrosis factor-alpha(TNF-alpha, interferon-alpha(IFN-alpha on human renal carcinoma cell line growth / 대한비뇨기과학회지

Recently much interest has been expressed for the use of biological response modifiers and growth factor antagonists in the treatment or radiation and chemotherapeutic drug-refractory renal cell carcinoma. Herein antiproliferative effects of Suramin, human recombinanl tumor necrosis factor alpha (TNF-alpha) and human recombinant interferon alpha (IFN-alpha) as a single and in combination were studied in vito on human renal carcinoma cell line (Caki-1). Antiproliferative effect was evaluated by trypan blue dye exclusion assay after 3 days exposure to Suramin at 10, 30, 100, 300, 1,000 mcg/ ml. TNF-alpha at 1, 10, 50, 100, 500, 1,000 units/ml and IFN-alpha at 10, 100, 1,000. 10,000 units/ml concentration, respectively. In addition, effects of combined administration in tolerable peak plasma level or suramin (300 mcg/ml). TNF-alpha (500 units/ml) and IFN-alpha (1000 units/ml) were comparatively analyzed to those of single drug administration. The results were as follows: 1. Significant dose dependent antiproliferative effects were shown by Suramin at above 100 mcg/ml. TNF-alpha at above 500 units/ml and IFN-alpha at above 10 units/ml, respectively (p<0.05). 2. At peak plasma level, suramin, TNF-alpha and IFN-alpha showed less than 50% inhibition of proliferation. 3. On combined administration, suramin plus TNF-alpha (61.0%), Suramin plus IFN-alpha(71.7%) and TNF-alpha plus IFN-alpha (57.2%) induced significantly greater inhibition of proliferation (p<0.005). These results suggest that further in vivo study using combination of Suramin plus TNF-alpha Suramin plus IFN-alpha and TNF-alpha plus IFN-alpha is necessary and that these combination trials may become one of the treatment options for renal cell carcinoma.