Local injection of OK-432/fibrinogen gel into head and neck carcinomas.

Immunotherapy with biological response modifiers (BRM) is a possible strategy against head and neck solid tumours. However, the rapid disappearance of BRM from the tumour area is one of the reasons for its limited clinical application. In this pilot study, fibrinogen gel containing OK-432 (a compound composed of attenuated Streptococcus pyogenes), an inducer of natural killer cells and T-cell cytotoxity, was injected directly into head and neck solid tumours of 15 patients. A dose of 5 Klinische Einheiten (KE) of OK-432 was reconstituted in 1 ml aprotinin and mixed with fibrinogen, the latter to maintain the OK-432 locally. 3 patients showed tumour regression, and in addition, we observed histological changes in the injected tumour of all patients. These results suggest that OK-432/fibrinogen gel generates a local immune response, leading to tumour regression.

Prediction of doxorubicin resistance in gastrointestinal cancer by P-glycoprotein staining.

Feasibility of immunohistochemical staining of P-glycoprotein for the prediction of doxorubicin resistance in gastrointestinal cancers was examined. Among 10 cancer cell lines which consist of two gastric cancer cell lines and eight colon cancer cell lines, seven cell lines were stained positively by the monoclonal antibody to P-glycoprotein, C219. In consequence of the evaluation on the effect of doxorubicin on these tumour cells by means of succinic dehydrogenase inhibition test (SDI test), zero out of seven cell lines stained positively by C219 was sensitive to doxorubicin, but two out of three cell lines stained negatively were sensitive. Among 23 fresh surgical specimens of gastrointestinal cancers which consisted of 15 gastric cancers and eight colon cancers, seven tumour tissues were stained positively by C219. All P-glycoprotein positive tumours were resistant to doxorubicin. On the other hand, four of 16 P-glycoprotein tumours were sensitive to doxorubicin. These data indicate that positively stained cancer cells by C219 are resistant to doxorubicin.

A case of ileus caused by a spiruroid nematode.

A 34-year-old male living in Aichi Prefecture, Japan, complained of lower abdominal pain. Ileus was suspected based on his clinical history and symptoms, and a laparotomy was performed. Four sections of a nematode were found in a large eosinophilic granuloma in the intestinal wall, and were identified as the larva of a spiruroid nematode. This is the third reported case of a spiruroid nematode infection found in the ileum.

Alteration of sensitivity to cis-diamine(glycolate)-platinum(II) (254-S) in oral tumor xenografts following multiple applications.

The emergence of resistance to platinum analogues is considered to be a major problem in the treatment of head and neck cancer. Therefore, it is important to clarify the mechanisms of resistance to these analogues and the mechanisms of the processes related to this resistance. The study of emergence of resistance in the solid tumors is particularly relevant. Ln the present study, the effect of a platinum analogue (254-S), on the response of an oral carcinoma cell line grown as a xenograft in nude mice, was studied. The effect of a full dose administered as a single intraperitoneal injection of 254-S (15 mg/kg X 1) on tumor growth was not significantly different from the effect of repeated intraperitoneal injections of 254-S, administered 3 times at 1/3 of this dose (5 mg/kg x3), or 5 times at 1/5 of this dose (3 mg/kg x5). However, when a single full-dose intraperitoneal injection of 254-S (15 mg/kg x1) was administered to each group of mice again at the 9th and 12th weeks after the initial treatment, different effects on tumor growth were observed among each group. The groups which received repeated treatment with 254-S (5 mg/kg, x3, or 3 mg/kg x5) showed a decrease in the inhibition of tumor growth, suggesting the emergence of resistance to 254-S. The study of platinum accumulation in the tumor tissues and a flow cytometric analysis of proliferating cell nuclear antigen (PCNA) supported the possibility that resistance to 254-S increases in tumor tissues treated repeatedly. These observations suggest that the potential use of this experimental assay as a model, may provide further insights into the therapeutic mechanisms of resistance to antineoplastic agents in the treatment of solid cancerous head and neck tumors.

Induction of apoptosis in head and neck tumor-cell lines by anti-fas antibody.

Programmed cell death, currently termed apoptosis plays a key role in the maintenance of the steady state in continuously renewing tissues. Since little is known of apoptosis in head and neck tumors, we studied morphological changes in head and neck tumor-derived cell lines (KB, KBrc, HSC-2,3,4), induced by anti-Fas antibody. Light and electron microscopic examinations were carried out after culturing these cell lines with the antibody for 1-2 days. The antitumor effect of anti-Fas antibody on tumor cells differed with the cell lines. Most of the cell lines that were sensitive to anti-Pas antibody showed evidence of enhanced apoptosis when the cells were pretreated with interferon-gamma. The results suggest that the strategy of induction of apoptosis by anti-Fas antibody may be considered in treatment of some tumors of head and neck.

Simultaneous xenotransplantation of human Graves' thyroid tissue and autologous bone marrow cells in severe combined immunodeficient mice: successful reconstitution of human Graves' hyperthyroidism.

Human thyroid xenografts and the autologous bone marrow (BM) cells from five patients with Graves' disease (GD) were simultaneously xenografted into severe combined immunodeficient (SCID) mice to study the role of BM cells for the perpetuation of human GD autoimmunity and hyperthyroidism. All SCID mice engrafted with thyroid tissue (TH) alone, TH + autologous peripheral blood mononuclear cells, and TH + autologous BM cells produced similar amounts of human IgG; however, the production in TH + BM-engrafted mice peaked later than that of mice without BM. Production of thyroperoxidase antibody and thyroglobulin antibody in TH + BM-bearing SCID mice peaked in later weeks after xenografting than in those without BM. Moreover, human Graves' hyperthyroidism was actually reconstituted in TH + BM-transplanted mice; this was confirmed by (A) significantly higher levels and longer periods of secreting thyroid-stimulating antibody than those in mice without BM engraftment. (B) persistent hyperthyroxinemia up to the end of the experiment. (C) extremely high radioidine uptake of the xenografted thyroid tissue, and (D) histological findings of the maintenance of hyperplastic change of the xenografted thyroid epithelial cells. Human BM stem cells (CD34) were identified only in mice with TH + BM xenografts when analyzed by immunohistochemistry. In conclusion, (A) we have developed an animal model for human hyperthyroid GD by simultaneous xenotransplantation of GD thyroid tissue plus autologous BM cells into SCID mice, and (B) BM cells have a crucial role for perpetuating human GD autoimmunity and hyperthyroidism in this system.

IFN-gamma has a protective role against thyroid-specific autoantibody production in severe combined immunodeficient (SCID) mice xenografted with Graves' thyroid tissue.

We studied the effects of exogenous human IFN-gamma or neutralizing monoclonal antibody (mAb) to IFN-gamma on xenografted human Graves' thyroid tissue in severe combined immunodeficient (SCID) mice to investigate a possible role of IFN-gamma in the pathogenesis of human Graves' disease. Human thyroid tissues from four patients with Graves' disease were xenografted into SCID mice. Two weeks after xenografting, mice were divided into three groups with human IgG levels similar to each other. Mice in the first group were treated with human IFN-gamma daily for 6 weeks; mice in the second (similar) group were treated with an mAb to IFN-gamma; mice in the third group were given mouse IgG only (control group). Blood samples were taken every 2 weeks for human IgG and thyroid-specific autoantibodies (Tg-Ab, TPO-Ab, and thyroid-stimulating antibody). After 6 weeks' treatment, mice were killed, and the thyroid xenograft was examined for thyrocyte HLA-DR expression. Human IgGs were produced equally in all three groups; mice treated with IFN-gamma showed significantly lower amounts of thyroid autoantibodies than those in the control group. Thyrocyte HLA-DR expression was markedly increased in xenografts from mice with IFN-gamma administration. On the other hand, anti-IFN-gamma mAb injection caused only slight suppression of HLA-DR expression on xenografted thyroid cells. In conclusion, IFN-gamma may down-regulate the production of thyroid-specific autoantibodies but not human IgG, at least under these circumstances; there thus may be specific inhibitory effects of IFN-gamma against thyroid-specific autoantibody production of intrathyroidal plasma cells, and this animal model may help to elucidate the possible role of cytokines in the pathogenesis of Graves' disease.

High urinary hydroxyproline excretion in patients with advanced head and neck cancer.

We studied the urinary excretion of hydroxyproline, measured as the hydroxyproline-to-creatinine ratio (HOP/Cr), in sixty-six patients with head and neck disease including, 18 patients with head and neck cancer. Urinary hydroxyproline excretion was determined before treatment by the linear gradient elution method. Although the urinary excretion of hydroxyproline is considered to be useful in monitoring cancer tissue such as prostate carcinomas, no studies have been performed on the urinary excretion of hydroxyproline in patients with head and neck cancer. The HOP/Cr was significantly higher in cancer patients than in the healthy control. The ratio of patients with T3/T4 cancers was higher than in patients with benign tumors (p<0.05), chronic inflammatory diseases (p<0.01) or T1/T2 cancer (p<0.05). The high HOP/Cr with T3/T4 head and neck cancer corresponded to the extent of tumor invasion into the surrounding tissues such as muscles and bone. This suggests that HOP/Cr may be useful as a supplementary parameter for the assessment of tumor invasiveness which is masked in head and neck carcinomas such as skull base or maxillary carcinomas.

An increase in laryngeal aerosol deposition by ultrasonic nebulizer therapy with intermittent vocalization.

In acute or chronic laryngitis as well as pharyngitis, nebulization with steroid and antibiotics is considered to be a safe and effective treatment. However, the optimal dose, frequency, duration, and method of administration by ultrasonic nebulization is not known. To evaluate whether there is continual improvement using ultrasonic nebulizer therapy, the deposition rate of aerosol particles into the upper airway was studied. We analyzed the deposition rate in the upper airway by a counting system of radioactive isotope (99mTc-diethylene triaminepentaacetic acid; 99mTc DTPA). Volunteers in good health inhaled an aerosol containing 99mTc DTPA under various conditions of inhalation. Deep and slow inhalation at a respiratory rate of 12 breaths/minute resulted in a high deposition rate of 99mTc DTPA in the lung and a low deposition rate in the larynx. In contrast, faster inhalation at a respiratory rate of 36 breaths/minute resulted in an increase in the isotope deposition rate in the larynx. Furthermore, when the volunteers vocalized intermittently during the fast inhalation at a respiratory rate of 36 breaths/minute, the isotope deposition rate in the volunteer's larynx was higher than in all other inhalation conditions. These results suggest that fast inhalation with intermittent vocalization is one of the important inhalation methods used to improve the deposition rate of aerosol particles into the upper airway with an ultrasonic nebulizer.

A simple method to estimate the secretion of saliva from minor salivary glands using iodine-starch reaction.

OBJECTIVE/HYPOTHESIS: This study was undertaken to detect the faculty of secretion of saliva from minor salivary glands by analyzing a color reaction on a test tape containing iodine and starch that was applied on the lower lip. STUDY DESIGN: A study involving 63 patients with oral dryness, 7 patients with Sjogren syndrome, and 70 healthy individuals was performed. METHODS: A test tape (1 x 1 cm) containing iodine and starch was set on the mucosal area anterior to the labia frenulum for 30 seconds. Because the number of blue spots was considered to correspond to the number of ostia of the salivary gland on the lower lip that was examined, the number of blue spots occurring as a reaction of iodine and starch on the test tape was counted and was compared among three groups. In addition, the relationship between the histopathological findings and the number of spots was analyzed. RESULTS: The average number of spots in the patients with oral dryness (4.52+/-3.18 [mean +/- SD]) was lower than that in healthy individuals (9.49+/-2.52, P <.01), and that in the patients with Sjögren syndrome (2.14+/-1.35) was the lowest among all groups in the study. Moreover, this reduction in the number of spots in those patients was accompanied by histopathological changes of the minor salivary glands. CONCLUSION: These findings suggest that this simple, noninvasive method can be successfully used for the estimation of the faculty of secretion of saliva from the minor salivary glands.

Laboratory maintenance of Eucyclops serrulatus (Copepoda: Cyclopoida).

A simple method for culturing a freshwater copepod Eucyclops serrulatus is described. A flagellate Chilomonas paramecium serves as the food organism. Starting from a single, egg-bearing female collected in a pond, the copepod proliferated for at least 10 generations apparently with no adverse changes. The stock culture has been maintained with minimal effort for at least 19 months. The method is expected to make the copepod useful in the laboratory as an intermediate host of various parasitic helminths.

[Mechanism of cisplatin and peplomycin therapy on head and neck carcinoma].

Results of chemotherapy for head and neck cancer are now improving owing to the development of concomitant use of chemotherapeutic agents such as cisplatin or peplomycin. Concomitant use of the two agents has been favored clinically. However, fundamental study on the combination therapy has not been carried out sufficiently. In the present study we studied the cell kinetics of tumor cells by the combination of cisplatin and peplomycin using flowcytometric analysis and electron microscopy. Survival of tumor cells was lowest, when peplomycin was administrated 3 days after cisplatin. Cell kinetics showed an accumulation at the S and G2M stage in this situation, By the electron microscopic study, microvilli of tumor cells disappeared and the blebbing of tumor cells was observed, when cisplatin alone was administrated. However, when the combination of cisplatin and peplomycin was administrated to the tumor cells, tumor cells enlarged. The results suggest that the mechanism of combination therapy differed from the mechanism of cisplatin alone.

A kinetic study of egg production, fecal egg output, and the rate of proglottid shedding in Hymenolepis nana.

To estimate the rate of shedding of the proglottids during normal development of Hymenolepis nana in mice, 2 parameters, i.e., egg production and fecal egg output, have been determined. Changes in the number of eggs/proglottid along the length of the worm showed that, not only in vitro but also in vivo, a considerable number of eggs is discharged in situ while the proglottids are still attached to the worm. The in situ egg discharge, however, accounts only for a small fraction of the eggs appearing in the feces, and it can be neglected in the estimation of the shedding of proglottids. In H. nana the larva rotates 90 degrees in the egg during its development. This seems to occur in a short period, quite in concert within each proglottid. The transition point (the proglottid where 50% of the eggs have begun rotating) can be a useful landmark to estimate the rate of proglottid shedding. Egg numbers at the transition points approximate the genuine egg production, because in situ egg discharge mostly occurs only after the proglottids have passed the transition point. The 3 processes, i.e., the rotation of the larva in the egg, the in situ egg discharge, and the shedding of proglottids, usually occur in an orderly way. However, on day 20 of infection, just before the worms are lost (or destrobilated), shedding of proglottids seems to be inhibited, whereas the other processes are apparently normal.(ABSTRACT TRUNCATED AT 250 WORDS)

Histamine H1 receptors and affinity analyses in human nasal mucosa in cases of nasal allergy.

An increased number (density) or sensitivity (affinity) of histamine H1 receptors could cause an increase of allergic symptoms. Using the specific H1 receptor blocker 3H-mepyramine, we have demonstrated for the first time an H1 receptor number (Bmax) and a binding affinity (Kd) in guinea pig and human nasal mucosa. H1 antihistamines inhibited that 3H-mepyramine binding sites represent H1 receptors in nasal mucosa. Bmax and Kd values were not significantly different among the nonallergic sinusitis, vasomotor rhinitis, or the nasal allergy group, suggesting that H1 receptors do not change quantitatively and qualitatively in allergic nasal mucosa. There was no significant correlation between H1 receptor number and clinical data (IgE, peripheral eosinophils, RAST). These data suggest that H1 receptor number and affinity are not main etiological and pathophysiological factors in nasal allergy.

Intracellular calcium changes and chemosensitivities of human epidermoid carcinoma cell lines after exposure to cisplatin.

In order to elucidate the mechanisms of cisplatin (cis-diamminedichloroplatinum; CDDP)-resistant tumor cells, we previously established a CDDP-resistant KB cell line (KBrc cells) from a parental KB cell line derived from epidermoid carcinoma (KB cells). The KBrc cells were resistant to 5 kinds of platinum (Pt) drugs. Intracellular Pt concentrations in KBrc cells were lower than in KB cells. Decrease of intracellular Pt concentrations was one of the CDDP-resistant mechanisms. When we measured changes of intracellular calcium ion concentration ([Ca2+]i) during exposure to high-dose CDDP, a sustained elevation of the [Ca2+]i level was observed in the KB cells. These results suggest that the mechanisms underlying CDDP resistance involve changes in calcium channels and an alteration of calcium homeostasis in the tumor cell line.

Effect of lymphokine-activated killer cells on head and neck tumours in nude mouse model.

The present study was designed to investigate the in vivo effect of local application of lymphokine-activated killer (LAK) cells on the growth of tumours implanted under the renal capsule in nude mice, and especially to test whether large granular lymphocytes (LGL), regarded as natural killer (NK) cells, are the main precursor of LAK cells in vivo. Our results showed that the local application of LAK cells inhibited the growth of tumours in the head and neck region. The growth of tumours implanted under the renal capsule was inhibited by local application of 1 x 10(7) recombinant interleukin-2 (rIL-2) activated non-adherent lymphocytes, but the inhibitory effect was almost the same as produced by 3 x 10(6) rIL-2-activated LGL application. The findings indicate that the rIL-2-activated LGL are the main effectors in inhibiting tumour growth. In addition, rIL-2-activated non-adherent lymphocytes as well as LGL significantly prolonged the number of days of 50% survival and mean survival time of nude mice, in which HLaC78 cells, from a laryngeal tumour cell line, were injected into the subrenal capsule space with effector cells at various effector: target (E:T) ratios. The results indicate that the application of LAK cells may be useful in the treatment of patients with head and neck tumours.

Elevation of cystathionine gamma-lyase activity in the serum of rats treated with a single dose of carbon tetrachloride.

Cystathionine gamma-lyase activity in the sera of rats subjected to experimental hepatotoxicity after intraperitoneal administration of carbon tetrachloride (CCl4) was measured and compared with activities of aspartate aminotransferase (GOT) and alanine aminotransferase (GPT), which have been clinically used for detecting liver damage. In the experimental subjects, serum levels of cystathionine gamma-lyase showed a similar behavior to GOT and GPT, increasing markedly with respect to the controls after administration of CCl4 and reaching a maximum at 24 hours. No such cystathionine gamma-lyase activity was detected immunochemically in the control subjects. These data suggest that measurement of serum cystathionine gamma-lyase activity could be used as a sensitive and specific marker of hepatic cytolysis.

[Establishing three dimensional tumors in vitro and the reaction of the lymphokine-activated killer cells (LAK) to the three dimensional tumors]. / Etablierung dreidimensionaler Tumoren in vitro und Wirkung von lymphokin-aktivierten Killer (LAK)-Zellen auf die dreidimensionalen Tumoren.

In this study, we attempted to develop a technique, by which three-dimensional tumors were produced from two cultured head and neck tumor cell lines (Hep2, KB) and a colon adenocarcinoma cell line (HT29) using fibrinogen, thrombin, and double layered agar system. The three-dimensional tumor was large enough to perform the histologic study, which showed no significant histologic difference in comparison with the histologic findings of the xenografted tumor on nude mice. Furthermore, we applied this assay model to evaluate the antitumor effect of lymphokine activated killer (LAK) cells on the three-dimensional tumor produced by the technique. When tumor cells were cocultured with LAK cells, the damage of the three-dimensional structure due to the degeneration of tumor cells was observed. These findings suggest that the three-dimensional tumor may be useful to evaluate the antitumor effect of LAK cells in term of head and neck solid tumors.

[Study on the preoperative adjuvant therapy of cancer--relation between serum and tumor tissue levels of UFT and OK-432 after administration, and skin reactions to Su-polysaccharide (Su-Ps)].

This study was carried out with 48 patients received surgery, i.e., 23 stomach cancer, 8 colon cancer, 6 rectal cancer, 9 breast cancer etc. Patients in group A received UFT in combination with OK-432. Each of UFT or OK-432 was given to the patients in groups B or C, respectively. Changes in the skin reaction to Su-PS were measured before and after dosing, and concentrations of Tegafur and 5-FU in serum and tumor tissues were determined after administration. Analysis of the skin reaction to Su-Ps revealed that patients with positive skin reaction before surgery in group A didn't manifest depression due to sensitization by UFT therapy. Although average values of the skin reaction after dosing were slightly lower compared to those before dosing in group B, sensitization was effective. Values of the skin reaction after dosing were significantly (p less than 0.05) high compared to those before dosing in groups A and C. Concentrations of Tegafur and 5-FU in serum reached to the peak 2 hr later and were maintained high enough to expect clinical responses even at 4 hr after administration in groups A and B. Especially there was not a significant difference between groups A and B in tumor tissue levels of 5-FU, and a high effective concentration was obtained. Combination therapy of UFT with OK-432 exhibited no significant interaction between them in adjuvant immuno-chemotherapy, and satisfactory results were expected in clinical cures.

[Study on the concentration of FT-207 and 5-FU in serum, lymph nodes and tissues after administration of FT-207 suppositories].

In 21 cancer patients receiving administration of FT-207 suppositories, we studied the concentration of FT-207 and 5-FU in the blood before surgery, and in the blood and tissues-tumor, normal and lymph node-sampled during surgery. These 21 cases were comprised of 12 cases of stomach cancer, 1 case of lymphosarcoma, 4 cases of rectal cancer, 2 cases of sigmoid cancer and 2 cases of breast cancer. Two hours after administration, the concentration of 5-FU reached its highest level and continued to maintain a level at which it could be expected to bring about clinical results even after four hours. The concentration of 5-FU in tumors and lymph nodes was high, especially in tumors compared with normal tissue (p less than 0.01). No difference could be found in the concentrations of FT-207 and 5-FU in serum and tissues from each 12 cases of stomach cancer, not only between of the early cancer and advanced cancer but also between patients more than 60 years old and those under 60 years old. No side-effects occurred in any case during suppository administration.