Partial Purification of Histaminase from Guinea-Pig Liver by Gel-Filtration at High Temperature

Histaminase was partially purified by Sephadex G-200 gel filtration at high temperature. Guinea pig liver histaminase was extracted with heparin. The extract was fractionated by Sephadex G-200 gel filtration at 4°C. In the fractions containing histaminase, 7 dense and 2 faint protein bands were detected on SDS-gel electrophoresis. Further, fractionation of this sample by Sephadex G-200 gel filtration at 55°C markedly decreased the number of bands, i.e. only 1 dense and 2 faint bands were observed in the fractions in which histaminase activity was detected, and the enzyme activity was increased by approximately ten times. The results suggest that gel filtration at high temperature may be useful for partial purification of histaminase.This work was supported by Scientific Research Grant from the Japanese Ministry of Education (B, No. 58480264)

The extract of syngeneic keratinocytes enhances IgE production from BALB/c mouse splenic lymphocytes in vitro.

Background: The increase of serum IgE levels is closely associated with atopic dermatitis. We have previously revealed that cellular extract of PAM212 cells (PAM-extract), BALB/c mouse keratinocyte cell line, induced a remarkable increase of serum IgE levels, in vivo, when subcutaneously injected into BALB/c mice. However, precise mechanism of IgE-increasing activity was unclear. Objective: To elucidate the mechanism of IgE-increase in sera of BALB/c mice induced by PAM-extract, we explored the direct influence of PAM-extract on immunoglobulin production and class-switching in the culture of splenic lymphocytes and purified B-cells, in vitro. Methods: Splenic lymphocytes or purified B-cells obtained from BALB/c mice were cultured with various combinations of IL-4, anti-CD40 antibody, and PAM-extract for seven days. IgE and IgG concentrations of culture supernatants were measured by ELISA. Epsilon germ-line transcriptions were assessed by RT-PCR from the cultured cells. Results: IgE and IgG concentrations in culture supernatant of splenic lymphocytes were increased by an addition of PAM-extract in the presence of both IL-4 and anti-CD40 antibody. Epsilon germ-line transcript was also induced in parallel to the increase of IgE production. Similar results were obtained when purified B-cells were employed in stead of whole splenic lymphocytes. Conclusion: The cellular extract of keratinocyte promotes immunoglobulin class-switching to IgE and IgE production from mouse splenic B-cells in an IL-4- and CD40-stimuli-dependent manner. Such enhancement may account for the increase of serum IgE in patients with dermatitis in association with a Th2 microenvironment

モルモット皮膚のヒスタミナーゼとその阻害因子

The crude histaminase preparation obtained from guinea pig skin was fractionated by Sephadex G-200 at 4°C and 55°C. The histaminase activity corresponded to the descending limb of the first peak of protein eluted just after void volume at 4°C and to the second peak at 55°C. The histaminase activity in the fractions eluted at 4°C was higher at 55°C than at 37°C. On the contrary, no significant difference between the histaminase activity assayed at 55°C and 37°C was observed in the fractions eluted at 55°C. On Sephadex G-200 gel filtration, histaminase was eluted in the later fractions at 55°C, compared with the fractions eluted at 4°C, indicating that the molecular weight of histaminase may become smaller at 55°C. These facts suggest that histaminase obtained from guniea pig skin may bind with an inhibitor-like substance at low temperature and that the enzyme may separate from the inhibitor at high temperature.This work was supported in part by Scientific Research Grant from the Japanese Ministry of Education (C, No. 56570383)

Therapeutic Guidelines for Atopic Dermatitis 2002

We summarized the Therapeutic Guidelines for Atopic Dermatitis 2002, and outlined important points with reference to medical care and management of atopic dermatitis

An Experimental Study on Occult Blood in Feces (Experiment by Oral Administration of Human Blood)

Examinations of occult blood in feces were performed after a certain amount of blood treated previously with hydrochloric acid limonnde was given into the stomach or duodenum through a Levin tube. The follwing results were obtained. 1. All the cases were positive in the reaction after the administration of more than 2 ml. of the blood. 2. No significant difference in the reaction between intragastric and intraduodenal administration. 3. The intensity of the positive reaction became weak as the acidity in gastric juice became high. 4. The longevity of the positive reaction was prolonged with larger amount of the blood given but they were not proportional when the amount of more than 3 ml. of the blood was administrated. After 4 days all the cases showed negative reaction