The molecular technique for 168 rRNA gene sequences analysis and Western Blot were applied for determination of the Bacillus probiotic strains used in biosubtyl production at IVAC. The results showed that the nucleotide sequence identity of the NT strains was 99.74% in comparison with B. pumilus and the identity of the DL strains was 99.61% in comparison with B. cereus strains from the GenBank/EMBL/DDBJ Database. The result of Western blot analysis showed that, the antigen components of the strains NT and DL were not similar in their antigens, and they were not similar to the B. subtilis ATCC6633.
Bacillus ; Blotting, Western

PURPOSE: To investigate the presence of adaptive response by low dose radiation in murine tumors in relation to radiation induced apoptosis as well as related mechanism. MATERIALS AND METHODS: Syngeneic murine tumors, OCa-I and HCa-I, were given 0.05 Gy pretreatment followed by therapeutic dose of 25 Gy radiation. Induction of apoptosis was analyzed for each treatment group. Regulating molecules of apoptosis, p53, Bcl-2, Bax, Bcl-X, were also analyzed by Western blotting. RESULTS: In 0.05 Gy pretreatment group of OCa-I, 25 Gy-induced apoptosis per 1000 cells was 229, which was estimated at 30% lower level than the expected (p<0.05). In contrast, this reduction in radiation induced apoptosis was not seen in HCa-I. In the expression of apoptosis regulating molecules, p53 increased in both tumors in response to radiation. Bcl-2 and Bax did not show significant change in both tumors however, the expression of Bcl-2 surpassed that of Bax in 0.05 Gy pretreatment group of OCa-I. Bcl-X was not expressed in OCa-I. In HCa-I, Bcl-X showed increased expression even with 0.05 Gy. CONCLUSION: Adaptive response by low dose radiation is shown in one murine tumor, OCa-I, in relation to radiation induced apoptosis. Apoptosis regulating molecules including Bcl-2/Bax and Bcl-X, appear to related. This study shows an evidence that adaptive response is present, but not a generalized phenomenon in vivo.
Apoptosis* ; Blotting, Western ; Radiation Dosage

BACKGROUND: Various kinds of hair products are widely used due to the increase of interest in hair styling. Cosmetic procedures such as permanent waving are very popular today, but the medical studies related to the meaning and restoration pattern of hair damage are mainly based on structural findings. In measuring the degree of hair damage by the moleculobiological methods rather than the structural studies, the findings seem to be highly objective and standardized. OBJECTIVE: The purpose of this study was to identify the patterns of hair damage and restoration through electrophoresis and western blot analysis of hair proteins. METHODS: The three volunteers who we selected as subjects did not have any specific medical illness and had not performed any special cosmetic procedures which could have caused hair damage during the six months before the study. We conducted permanent waving on them. Human hair samples were obtained from the occipital scalp, which were that was not affected by the androgen. We performed extraction and concentration of the whole and partial hair protein, then operated electrophoresis and western blot analysis of the hair protein. RESULTS: In the western blot analysis of whole hair proteins, there was one positive finding on subject A. This may have resulted from the small amount of partial proteins among the whole hair proteins. In the western blot analysis of partial hair proteins, subject A and B showed positive findings. In particular, positive findings were found on the 14th week of the experiment. CONCLUSION: These results show a change in the hair proteins due to hair damage, and ultrastructurally, we found the possibility of prologation of actual hair damage longer than expected.
Blotting, Western ; Electrophoresis ; Hair* ; Humans ; Scalp ; Volunteers

OBJECTIVE: Survivin is an inhibitor of apoptosis protein(IAP), which inhibits apoptosis through a pathway distinct from the Bcl-2 family members. Overexpression of survivin and Bcl-2 have been commonly reported in human neoplasms. The authors investigate whether there is a synergistic effect on the anti-apoptosis rate of primary brain tumors "in situ" based on the co-expression of survivin and Bcl-2. METHODS: One hundred and two brain tumor patients who had been resected were included in this study. Survivin and Bcl-2 were detected by Western blotting analysis, while apoptosis was examined by DNA fragmentation analysis. An anti-apoptotic rate was assessed in these brain tumor samples based on the expression of survivin and Bcl-2 or co-expression of both. RESULTS: Survivin and Bcl-2 were expressed in 57(55.9%) and 53(52.0%) of 102 brain tumor samples studied respectively, and co-expressed in 31(30.4%). The percentage of astrocytic and meningeal tumors expressing survivin was significantly correlated with histological grades; however, Bcl-2 was not correlated (p=0.106). The anti-apoptotic rate in primary brain tumors with survivin, Bcl-2, and both was detected in 49(86.0%) of 57 samples, 42(79.9%) of 53 samples, and 27(87.1%) of 31 samples, respectively. Their difference in the frequency of anti-apoptosis was not significant. CONCLUSION: Survivin or Bcl-2 is involved in the anti-apoptosis. However, it suggests that co-expression of survivin and Bcl-2, together, have no synergistic effect on the anti-apoptotic properties of the primary brain tumors.
Apoptosis ; Blotting, Western ; Brain Neoplasms* ; DNA Fragmentation ; Humans ; Meningeal Neoplasms

BACKGROUND: It has been reported that immunological mechanisms may play an important role in the pathogenesis of vitiligo. Cellular degeneration in vitiligo is not limited to melanocytes but includes keratinocytes and probably whole epidermal melanin units. OBJECTIVE: The purpose of this study was to demonstrate the presence of anti-keratinocyte antibodies and the relationship between the level of antibodies and activity of the disease. METHODS: We analyzed the level of anti-keratinocyte antibodies in 13 patients with non-active, segmental vitiligo and 7 patients with active, non-segmental vitiligo by Western blotting. RESULTS: Using the Western blot method, we demonstrated the occurrence of anti-keratinocyte antibodies with keratinocyte cytoplasm. Anti-keratinocyte antibodies were more reactive in patients with active, non-segmental vitiligo than in those with non-active, segmental vitiligo. CONCLUSION: These results may be a secondary phenomenon to keratinocyte damage with concurrent occurrence of melanocyte destruction. However, the level of these antibodies may represent the acitivity of vitiligo.
Antibodies* ; Blotting, Western* ; Cytoplasm ; Humans* ; Keratinocytes* ; Melanins ; Melanocytes ; Vitiligo*

OBJECTIVETo express and purify recombinant human interleukin-35[IL-35-IgG1 (Fc) in eukaryotic expression system and to study the interaction of IL-35 with gp130 protein.

METHODSA mammalian expression vector, pSTEP2-IL35-LFc, was constructed and transfected into HEK293T cells. Then rhIL-35-IgG1 (Fc) was expressed and purified with protein A affinity chromatography, and was examined with SDS-PAGE and Western blot analysis. The binding of IL-35 to its receptor gp130 was investigated using enzyme-linked immunosorbent assay (ELISA). The biological effect of IL-35 on gp130 was explored in M1 myeloid leukemia cells.

RESULTSrhIL-35-Fc with high purity on reduced SDS-PAGE was obtained. ELISA confirmed that IL-35-Fc was bound to gp130 and neutralized the function of gp130 in M1 myeloid leukemic cells.

CONCLUSIONSHigh-purity and biologically active rhIL-35-Fc protein successfully produced in this study. IL-35 binds to gp130 and neutralizes its activity of in M1 myeloid leukemic cells.

Since CKD-712 has been developed as an anti-inflammatory agent, we examined the effect of CKD-712 during TLR4 signaling. Using HEK293 cells expressing TLR4, CKD-712 was pre-treated 1 hr before LPS stimulation. Activation of NF-kappaB was assessed by promoter assay. The activation of ERK, JNK, p38, IRF3 and Akt was measured by western blotting. CKD-712 inhibited the NF-kappaB signaling triggered by LPS. The activation of ERK, JNK, p38 or IRF3 was not inhibited by CKD-712. On the contrary the activation of these molecules was augmented slightly. The activation of Akt with stimulation of LPS was also enhanced with CKD-712 pre-treatment at lower concentration, but was inhibited at higher concentration. We suggest that during TLR4 signaling CKD-712 inhibits NF-kappaB activation. However, CKD-712 augmented the activation of Akt as well as Map kinases. Therefore, we suggest that CKD-712 might have a role as an immunomodulator.
Blotting, Western ; HEK293 Cells ; NF-kappa B ; Phosphotransferases ; Tetrahydroisoquinolines

The present study investigates the role of TGF-beta1 as one of possible etiologic factors of glaucoma by studying the effect of TGF-beta1 on the secretion of matrix metalloproteinase and tissue inhibitor of matrix metalloproteinase. The cultured samples of trabecular meshwork cells were treated with TGF-beta1 in concentrations of 0, 100, 1000 and 10000 pg/ml. The samples were analysed for the secretion of MMP2 and timp2 through electrophoresis, western blot and TGF-beta1 increased, the secretions of MMP2 and TIMP2 were not significantly changed after 24 hours. But the secretion of MMP2 was decreased while the secretion of TIMP2 was increased after 72 hours. The results suggest that TGFbeta1 may be one of the etilogic factors of glaucoma.
Blotting, Western ; Electrophoresis ; Glaucoma ; Trabecular Meshwork ; Transforming Growth Factor beta1

OBJECTIVE: The purpose of this study was to identify the ATP-sensitive potassium (K(ATP)) channel subtypes in uterine leiomyoma and normal myometrial cells, and to compare the difference in its expression between uterine leiomyoma and normal myometrial cells. METHODS: Fresh ten uterine leiomyomas and their adjacent normal myometrial tissues were obtained from hysterectomies that were conducted on benign diseases. With the use of reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis, we analysed the expression of K(ATP) channel subtypes in tissues and primary cultured leiomyoma cells. To demonstrate the K(ATP) channel activity in uterine leiomyoma cells, whole cell patch-clamp recordings were made. RESULTS: The uterine leiomyoma and normal myometrial tissues contained Kir6.1/SUR2B and Kir6.2/ SUR2B mRNAs, although the expression of SUR1 and SUR2A mRNAs were not expressed in the uterine leiomyoma and normal myometrial tissues. Primary cultured uterine leiomyoma and normal myometrial cells demonstrated same patterns. To determine the expression levels of K(ATP) channel subunits, high levels of Kir6.1, Kir6.2, and SUR2B were detected by western-blot analysis in uterine leiomyoma compared with normal myometrial tissues. K(ATP) channel currents were increased by estrogen application in uterine leiomyoma cells. CONCLUSION: These studies provide new knowledge concerning K(ATP) channels in uterine leiomyoma and normal myomtrial cells. we demonstrated uterine leiomyoma and normal myometrial tissues contained Kir6.1/ SUR2B and Kir6.2/SUR2B subunits and showed an increased expression in uterine leiomyoma compared with normal myometrial tissues. These results suggest that K(ATP) channels are important elements in growth of uterine leiomyoma.
Blotting, Western ; Estrogens ; Hysterectomy ; Leiomyoma* ; Potassium ; RNA, Messenger

At the level of individual cells, signaling is crucial in cell division, differentiation, metabolic control and death. Reception of the signals depends on receptor proteins that are usually at the cell surface, and these receptor proteins bind the signal molecule. The binding activates the receptor, which in turn activates one or more of the intra-cellular signaling pathways. These relay chains of molecules, mainly intra-cellular signaling proteins, process the signal inside the receiving cell and distribute it to the appropriate intra-cellular targets. Cell signaling pathways are involved in the pathophysiology of many diseases and also in the mechanisms of action of many drugs, including local and general anesthetics. Knowledge of the basic cell signaling mechanisms is essential for understanding many of the pathophysiologic and pharmacologic mechanisms. Therefore, if we focus on applying the new cellular and molecular biologic research, these efforts could identify the mechanism of diseases and help develop new drugs in the field of anesthesiology and pain medicine.
Anesthesiology ; Anesthetics, General ; Blotting, Western ; Cell Division ; Proteins