Coffee (Coffea spp.) is one of the most important agricultural commodities, being widely consumed in the world. Various beneficial health effects of coffee have been extensively investigated, but data on habitual coffee consumption and its bio-physiological effect have not been clearly explained as well as it is not proved the cause and effect between drinking coffee and its bio-physiological reactions. We made the dialyzed coffee extract (DCE), which is absorbable through gastrointestinal tract, in order to elucidate the cellular effect of whole small coffee molecules. RAW 264.7 cells, a murine macrophage lineage, were directly treated with DCE, i.e., DCE-2.5 (equivalent to 2.5 cups of coffee a day), DCE-5, and DCE-10, for 12 hours, and their protein extracts were examined by immunoprecipitation high performance liquid chromatography (IP-HPLC). RAW 264.7 cells differently expressed the inflammation-related proteins depending on the doses of DCE. RAW 264.7 cells treated with DCE showed marked increase of cathepsin C, cathepsin G, CD20, CD28, CD31, CD68, indicating the activation of innate immunity. Particularly, the macrophage biomarkers, cathepsin G, cathepsin C, CD31, and CD68 were markedly increased after DCE-5 and DCE-10 treatments, and the lymphocyte biomarkers, CD20 and CD28 were consistently increased and became marked after DCE-10 treatment. On the other hand, RAW 264.7 cells treated with DCE showed consistent increase of IL-10, an anti-inflammatory factor, but gradual decreases of different pro-inflammatory proteins including TNFα, COX-2, lysozyme, MMP-2, and MMP-3. In particular, the cellular signaling of inflammation was gradually mitigated by the reduction of TNFα, COX-2, IL-12, and M-CSF, and also the matrix inflammatory reaction was reduced by marked deceases of MMP-2, MMP-3, and lysozyme. These anti-inflammatory expressions were consistently found until DCE-10 treatment. Therefore, it is presumed that DCE may have dynamic effects of innate immunity activation and pro-inflammation suppression on RAW264.7 cells simultaneously. These effects were consistently found in the highest dose of coffee, DCE-10 (equivalent to 10 cups of coffee a day in man), that might imply the small coffee molecules were accumulated in RAW 264.7 cells after DCE-10 treatment and produce synergistic cytokine effects for innate immunity activation and anti-inflammatory reaction concurrently.

Coffee is one of the most familiar beverages to modern human adults, but its bio-physiological effect has not been clearly elucidated. It was known that more than one thousand chemicals were included in the ordinary coffee extract. Among them, the caffein and chlorogenic acid (caffeoylquinic acids) are most abundant and have been investigated by many authors so far. In order to know the real cellular effect of whole coffee extract elements, the dialyzed coffee extract (DCE)1) was made to get coffee elements less than 1000 Da molecular weight, which are freely absorable through gastrointestinal tract. It was directly treated in the culture of RAW 264.7 cells, a murine macrophage lineage. RAW 264.7 cells were treated with DCE equivalent to 2.5 cups of coffee (DCE-2.5), DCE-5, and DCE-10 for 12 hours, and their protein extracts were examined by histological observation and immunoprecipitation high performance liquid chromatography (IP-HPLC). RAW 264.7 cells differently expressed the proliferation-related proteins depending on the dose of DCE. DCE-2.5 and DCE-5 enhanced the cellular growth of RAW 264.7 cells by increasing the expression of β-actin, PCNA, Ki-67, MPM2, MAX, cMyc, E2F-1, and Rb-1, and by decreasing the expression of MAD and p21. These proliferation-related proteins were rarely affected by DCE-10. DCE-2.5 and DCE-5 induced the cellular proliferation of RAW 264.7 cells by the signaling of E2F-1 and cMyc, respectively, but these cellular effects almost disappeared in DCE-10. Therefore, it was presumed that the low dose of coffee, DCE-2.5 and DCE-5 might be effective for the proliferation of murine macrophages, RAW264.7 cells, contrast to the high dose of coffee, DCE-10. It was also suggested that the low dose of DCE-2.5 and DCE-5 be helpful to increase the innate immunity in vivo by increasing the cell number of macrophages in contrast to the high dose of DCE-10.

In order to perform the biological investigation of coffee extract containing different molecules, it would be necessary to develop in vitro experimental system rather than animal experiment. Although the animal experiment treated via oral intake or intravenous injection may disclose the whole systemic effect, the in vitro cell culture experiment would be more convenient to analyze direct cellular effect of coffee extract than animal experiment. Therefore, this study was aimed to develop a dialysis method for the crude coffee extract to perform the biological investigation using murine macrophage cell line, RAW 264.7. First of all, the RAW 264.7 cells treated with dialyzed coffee extract were observed, and subsequently their protein extracts were analyzed by gel filtration chromatography, thin layer chromatography, and immunoprecipitation high performance liquid chromatography (IP-HPLC). Resultantly, it was found that the low dose (20μg/mL) of dialyzed coffee extract, about 5 cups of ordinary coffee drinking for human adult, enhanced the growth of RAW 264.7 cells by increased expression of β-actin and Ki-67, and also induced the anti-inflammatory effect by decreased expression of NFkB, TNFα, and LC3 contrast to the high dose (40μg/mL) of dialyzed coffee extract. The low dose of dialyzed coffee extract produced almost no harmful effect on RAW cell culture for 12 hours, rather than it produced stimulatory effect on RAW cells by increasing the cell number and enhancing the protein expression of β-actin, Ki-67. Therefore, it was thought that the low dose of dialyzed coffee extract is applicable to cell culture experiment without difficult purification procedures of coffee elements. In addition, as the contrast cellular effect between the low and high dose of coffee extract was found in this study, it was also presumed that the low dose of coffee extract may play an important role in the inflammatory reaction of murine macrophages.

The purpose of this study was to investigate the distribution of trig˗ ger points(TrPs) on athletes with various sporting background. To achieve the purpose, a study was carried out through a survey from 180 athletes involved in 6 selected sports at Yong-In University. Selected sports included Judo, Taekwondo, Kendo, Ssi-reum(Korean traditional wrestling), Boxing, and Golf. An interview type survey and physical examination were conducted with each thirty athletes from each of the selected sports groups. Technical statistic(SPSS 15.0) was used to analyze the distribution of TrPs on these athletes. The most common TrPs observed in muscles of Trapezius, Quadratus Lumborum, Quadriceps in Judo. In Taekwondo, it was on the trapez˗ ius and triceps surae. Kendo athletes had TrPs at sites of trapezius, brachioradialis and triceps surae. Ssirem athletes were found to have TrPs on trapezius, deltoid and quadrates lumborum. In boxers, TrPs appearing at trapezius and brachioradialis were observed. Finally, Golf players were seen to have TrPs at trapezius, quadrates lumbo˗ rum and brachioradialis. Hence, the analysis shows that there are significant differences of the distribution of TrPs according to the different sport items of the athlete.

In order to perform the protein analysis using the paraffin sections previous fixed with formalin, we applied the ImmunoMemBlot (IMB) method1) to detect the epitopes of target proteins with specific antibodies. In this study the protein extracts were obtained from the paraffin sections of each representative case of ameloblastoma, adenomatoid odontogenic tumor (AOT), and normal gingiva, and more a protein extract from fresh tissue of ameloblastoma was also compared to evaluate the IMB results used with 24 different antibodies. First of all, in the comparison between the paraffin section extract and fresh tissue extract of ameloblastoma, the latter consistently showed more positive IMB reaction than the former. Meanwhile, the paraffin section extract of ameloblastoma was more comparable with that of normal gingival, disclosing that most of proliferating genes, oncogenes, and apoptosis related genes, i.e., PCNA, CDK4, c-erbB2, CEA, p53, Bax, Bad, FLIP, FAS, Bcl-2, p21, N-ras, MMP-2, MMP-9, caspase-3, -8, -9, were highly expressed in ameloblastoma, but EGFR, HGF, and VEGF were similarly expressed both in the ameloblastoma and in normal human gingiva. On the other hand, the comparison between ameloblastoma and AOT both in the immunohistochemistry and IMB using their paraffin section extracts clearly demonstrated that the ameloblastoma showed more expression of proliferating genes and oncogenes while the AOT showed more expression of apoptosis related genes, i.e., Bax, Bad, FLIP, and caspase-9. Taken together, these data suggest that the IMB can be used for the primary screening of quantitative protein analysis using the paraffin section extract, and that the IMB results could be evaluated in conjunction with the immunohistochemical observation.