Tatsuhito Tono-Oka

Chemiluminescence of whole blood: 1. A simple and rapid method for the estimation of phagocytic function of granulocytes and opsonic activity in whole blood

Luminol-dependent chemiluminescence of whole blood (whole blood CL) was developed to estimate the phagocytic function of granulocytes and the serum opsonic activity simultaneously. Whole blood (0.1 ml) was examined directly and results were obtained within 20 minutes. Phagocytic function of granulocytes can be estimated from the peak CL of whole blood and the number of granulocytes in a specimen, and the opsonic activity from the amount of time the peak CL is shown after the addition of nonopsonized CL inducer. Thus this method, using several types of CL inducer, offers much information concerning the functions of the phagocytic system in whole blood.

Chemiluminescence of whole blood: II. Application to clinical examination of phagocytic functions of whole blood from various types of disease

Measurements of whole blood chemiluminescence (CL) were carried out by using 0.1 ml of blood to evaluate phagocytic functions from various kinds of pediatric patients. Patients with definite bacterial infections showed an elevated background CL and high peak CL per 1000 granulocytes after stimulation with zymosan. These results suggest that granulocytes of these patients had been preactivated in vivo, their phagocytic and/or metabolic activity being enhanced. Contrary to these results, blood from the patients with viral infections showed almost no abnormalities. Patients with chronic granulomatous disease showed no significant CL by this method similar to conventional ones. This method was also shown to be a useful technique for monitoring the phagocytic functions of blood after granulocyte transfusion.

Characteristics of Impaired Chemotactic Function in Cord Blood Leukocytes

Summary: Mobilities of cord blood granulocytes were studied using the agarose plate method and Boydens chamber method. In the agarose plate, granulocytes of cord blood were shown to have moderately decreased responses in chemotaxis and chemokinesis induced by Escherichia coli-derived chemotactic factor and/or zymosan-activated serum, whereas they were shown to have a normal capacity of random mobility. Although their distance and index of chemotaxis or chemokinesis in the agarose plate were significantly less than those of adult granulocytes, response rate in both types of mobility were evidently higher compared with those in patients with chemotactic defect. Furthermore, there is a difference between chemotactic responses of cord blood granulocytes to E. coli-derived chemotactic factor and to zymosan-activated serum in the agarose plate method. Using the latter, a more distinguishable difference between chemotactic responses of cord blood granulocytes and adult granulocytes was shown.The Boydens chamber method tended to show a more significant difference between chemotactic responses of granulocytes of cord blood and adults than in the agarose plate method.Speculation: The impaired chemotactic function of the cord blood granulocyte may be due to developmental immaturity in chemokinetic and/or chemotactic response, and the degree of impairment may be moderate.

Evaluation of the proliferative response of lymphocytes by measurement of intracellular ATP

The measurement of intracellular ATP levels by luciferin-luciferase-induced bioluminescence was used for the evaluation of proliferative responses of human mononuclear cells to lectins. A linear relationship was observed between the number of cells and the amount of ATP in the samples, and high reproducibility was obtained. The ATP content of samples containing lectin-stimulated mononuclear cells increased with time, and a significant difference from unstimulated cells was obtained 48 h after initiation of the culture. Furthermore pretreatment with mitomycin C strikingly increased the ATP level of lectin-stimulated cells, but not that of unstimulated cells was obtained. These findings suggest that the proliferative response of mononuclear cells can be measured simply without the use of isotopes, and that earlier events occurring in stimulated cells may be analyzed by this simple method.

A Case of Yersinia pseudotuberculosis Septicemia Accompanied by a Large Abdominal Tumor

A 2-year-old girl with Yersinia pseudotuberculosis septicemia had a large abdominal mass that had to be differentiated from malignant tumor. The mass disappeared rapidly with antibiotic therapy and was defined to be a cluster of enlarged ileocecal lymph nodes by the clinical course and the findings at ultrasonic examination and Gallium scintigraphy.

Enhanced granulocyte mobility induced by chemotactic factor in the agarose plate.

Summary Human granulocyte mobility under various conditions of chemotactic stimulus was studied using the agarose plate method. Enhanced mobility was observed when granulocytes were incubated in the agarose plate containing chemotactic factor generated from E. coli. A dose response type relationship was observed between the degree of enhanced mobility and the concentrations of chemotactic factor in a range of less than 10%. The rate of mobility was rapid up to 3 hr, after which time it was very slow. Preincubation of granulocytes with chemotactic factor of various concentrations did not have any influence on granulocyte mobility assayed after preincubation. The degree of mobility tends to be determined by the final concentration of chemotactic factor coming in contact with granulocytes. Thus granulocytes under a negative concentration gradient also showed an enhanced mobility. On the basis of these findings, we propose the hypothesis that the accumulation of granulocytes at the site of inflammation can be in part explained by chemokinesis, i.e. enhanced random mobility.

Whole Blood Chemiluminescence As an Acute Phase Reactant

Chemiluminescent response of whole blood was estimated as a function of acute phase reactant and compared with ESR and CRP in various types of pediatric patients with fever. 0.1 ml of whole blood was used as the material, and two parameters concerning the phagocytic function of whole blood were used: (1) Peak CL; total phagocytic function of whole blood reflecting the number of granulocytes in a specimen and their function; (2) Peak CL per 103 whereas viral infections consisting of upper respiratory infection, varicella and systemic bacterial infections showed an increased peak CL, not only due to the increased number of granulocytes, but also due to the enhanced CL response of the granulocytes themselves (thus peak CL per 103 PMN was laso increased), whereas viral infections consisting of upper respiratory infection, varicella and mumps, and acute leukemia in the active stage did not have a remarkable influence on the two parameters. In many types of febrile illness peak CL/103 PMN was significantly correlated with ESR and CRP, but not with granulocyte counts. ESR and CRP also did not correlate with granulocyte counts. Thus it was confirmed that the whole blood CL method is useful as a quick test with a small amount of specimen for evaluating febrile patients; concerning the intensity of inflammation as well as phagocyte (granulocyte) functions.

A Case of Mixed Bacterial Meningitis Associated with a Congenital Spinal Epidermoid

A case of mixed bacterial meningitis with congenital spinal epidermoid is reported. E. coli and S. faecalis were isolated simultaneously from the cerebrospinal fluid of a 14‐month‐old girl who had a continuing fever for 5 days. The clinical response was very poor, whereas bacteria detected showed high sensitivity in vitro to the antibiotics used clinically. Thus a myelography and a CT scanning of the lumbosacral region were performed one month after admission, when a spinal epidermoid was forced, containing an abscess connected to a thin dermal sinus which could not be recognized macroscopically. The patient recovered completely after surgical treatment.

Chromosomal Analysis of Childhood Acute Lymphoblastic Leukemia: As a Routine Examination for Diagnosis and Prognosis

Chromosomal analysis was performed as a routine examination for diagnostic and prognostic evaluation of children with acute lymphoblastic leukemia (ALL) in six cases encountered during a one‐year period. The results obtained are as follows. 1. Chromosomal abnormality of leukemic cells was observed in five out of six cases (approximately 80%). 2. Translocation 4; 11 or 14q+, which are known as a risk factor of ALL, were observed in three patients. Two of three patients died within six months of the onset of disease. One case was diagnosed as congenital leukemia with remarkable leukocytosis, and the other case was accompanied by hypereosinophilic syndrome. The remaining one patient, who is now in complete remission, is 13 years old, which is within the period of risk ages exceeding 10 years of age. 3. The case with 1q+ had no risk factor; however, he had a relapse 19 months after the diagnosis. Thus this particular chromosomal rearrangement appeared to be one of the risk factors. 4. The case with 6q‐, which has been reported to have a good prognosis in some cases of ALL, has no risk factor, and has been in complete remission. These results seem to confirm the usefulness of chromosomal analysis for the evaluation of the clinical course of ALL.

Study on Leukocytes Mobility in Pediatric Malignant Diseases during Chemotherapy

Mobilities of granulocytes and monocytes in 17 patients with infantile malignant diseases were estimated using the agarose plate method. Diseases were acute leukemia, neuroblastoma and rhabdomyosarcoma. All the patients were under the chemotherapy with multiple drugs for the remission induction or maintenance. Monocytes of 10 of 13 patients showed decreased random mobility, whereas that of granulocytes of all the patients was within a normal range. Chemotactic responses of monocytes to zymosan activated serum tended to be impaired in 5 of 11 patients, and that of granulocytes tended to be impaired in 12 of 20 patients. On the other hand, chemotactic responses of granulocytes to Escherichia coliderived factor did not tend to be impaired. This dissociation of chemotactic responses suggested that impaired chemotactic responses of granulocytes was factor specific. All these impairments were thought to be caused by chemotherapeutic agents, because there were no differences in the degree of the impairment between the groups in remission with maintenance chemotherapy and in an active stage with induction chemotherapy. From these results we speculated that the impaired chemotactic responses of granulocytes may partly cause the vulnerability to infections of patients with a malignant disease, and that the impaired random mobility and chemotactic responses of monocytes may cause partly impaired cellular immunity in these patients.