Lack of leukocyte migration inhibition by hepatitis B surface antigen in hepatocellular carcinoma patients.

Soluble part of hepatocellular carcinoma (HCC) tissue extracts with or without hepatitis B surface antigen (HBsAg) was tested against leukocytes of 13 histologically confirmed HCC patients. Inhibition of leukocyte migration was observed in 9 out of 13 cases when tested by soluble HCC extract containing HBsAg, while inhibition of lukocyte migration was observed in 8 out of 13 cases when tested by solublp greater than 0.05, by Fisher's exact test). In the meantime, soluble HCC extract with or without HBsAg did not significantly cause inhibition of leukocyte migration in 12 non-HCC patients. Therefore, it is concluded that inhibition of leukocyte migration in HCC patients is caused by the tumor-associated antigen, not caused by HBsAg.     

Effect of serum fractions from cancer patients on leukocyte migration out of capillary tubes]

Blood serum of patients suffering from cancer of the stomach and urinary bladder inhibited in vitro migration of autologous leukocytes, leukocytes of donors and control patients, and also guinea pig macrophages in over half of cases. In chromatography of these sera on Sephadex G-100 the activity inhibiting the leukocyte migration was revealed in fraction I (Mol. wt. over 100000) and in fractions IV and V (Mol. wt under 30 000). The blood serum and its fractions from cancer patients failed to eliminate the leukocyte migration inhibition caused by the tumour antigens in comparison with the leukocyte migration in the medium with control serum without any antigens. As suggested, the activity of fraction I inhibiting the leukocyte migration was due to the antigen-antibody complex, and of fraction IV and V--to a factor similar by its properties to the factor produced in vitro by lymphocytes stimulated by the antigens or mitogens.     

Leukocyte migration inhibition test (LMIT) in systemic lupus erythematosus.

A leukocyte migration inhibition test (LMIT) utilizing the agarose gel technique was performed with native DNA as an antigen in ten patients with systemic lupus erythematosus (SLE) and five normal subjects. Irrespective of disease activity, supernatants obtained at different time intervals during lymphocyte culture in eight patients with SLE showed significant alteration of migration, either enhancement or inhibition, of normal leukocytes. However, supernatants in the control experiments produced no significant alteration of migration. Polyacrylamide gel electrophoresis of supernatants obtained from the SLE group revealed that the inhibitory activity was present in the albumin region, whereas the enhancement activity was found in the beta-globulin region. These results indicate that the hitherto employed estimation of the leukocyte migration inhibition test based on the total activity of these two factors is insufficient for accurate evaluation of chemical mediators from sensitized lymphocytes and that the separation of these two factors may be important for a greater understanding of cellular immunity.     

Interrelationships in experiments in vitro between the migration activity of leukocytes and RNA and DNA synthesis]

The following correlations were revealed in the parallel study of leukocyte migration in vitro in the presence of a specific antigen and of spontaneous RNA and DNA synthesis in the cultured lymphocytes: 1) a direct correlation between the RNA and DNA synthesis in lymphocytes; 2) a close correlation between the antigen-induced migration and the levels of RNA and DNA synthesis. The effect of the antigen was evidenced by the inhibition or stimulation of leukocyte migration. A high ratio of RNA synthesis to DNA synthesis corresponded to the migration inhibition and a low one--to the migration stimulation. The ratio value varied mainly on account of the changes in the level of DNA synthesis. Participation of T and B cells in the regulation of the antigen-induced leukocyte mobility is discussed.     

Cross-reactivity of lung cancer patients' leucocytes in the leucocyte migration inhibition test

Summary Panels of 3 M KCl extracts of squamous-cell carcinomas, adenocarcinomas and oat-cell carcinomas of the lung were used for a comprehensive analysis of cross-reactivity in the leucocyte migration test. Lung cancer patients' leucocytes showed positive reactivity in 69%–100% of cases (n=353). No significant differences were observed when data were grouped with respect to the histological type of the tumours used for extraction or of the tumours of the leukocyte donors. Leukocytes of patients bearing tumours of nonpulmonary origin exposed to lung cancer extract panels and leukocytes of lung cancer patients exposed to gastrointestinal cancer extract panels were definitely less reactive (35%–47% and 6%–38%, respectively). However, a high reaction frequency was found in patients with lung metastases from different nonpulmonary tumours. This group of patients also frequently showed reactivity (52%) with normal lung tissue extracts. Patients with benign lung diseases reacted positively with lung tumour extracts in 25%–39% of cases, but donors with other benign disease and healthy controls were virtually nonreactive (0–14%).Hence, a high degree of cross-reactivity occurs in the lung cancer system and restricted cross-reactivity occurs with tumours of other organs. Possible explanations for the lung-oriented reactivity of patients with lung metastases are discussed.Abbreviations LMI leucocyte migration inhibition - MI migration index - LMT leucocyte migration test - SCC squamous-cell carcinoma - OCC oat-cell carcinoma - AC adenocarcinoma     

Leukocyte migration inhibition test in breast cancer patients treated with conventional therapy or with conventional therapy plus poly A · poly U

Summary Leukocyte migration inhibition tests (LMIT) were performed to measure the immunologic reactivity of patients with initially operable breast cancer. Leukocyte migration was inhibited in 38% of 159 patients in the presence of autologous tumor extract, in 33% of 160 subjects in the presence of autologous serum, and in 47% of 148 patients in the presence of extract and serum. These patients formed part of a randomized clinical trial comparing conventional treatment complemented by injections of poly A · poly U with conventional treatment and injection of saline as a placebo.A sequential study was carried out, testing the leukocyte reactivity 7 days, 2 months, 4 months, and 1 year after the operation. The percentage of patients with positive LMIT increased significantly with time. Statistical comparison revealed no significant difference in the reaction of the two therapeutic groups. In addition, no significant difference was found between those with lymph node involvement and those without, although the percentage of positive LMIT in the presence of tumor extract appeared to be higher in the second group 1 year after surgery. Radiotherapy given to those with lymph node involvement did not significantly change their reaction.This study also confirmed the presence in some autologous sera of a synergistic factor (SS factor) that increased the inhibition of migration of leukocytes by autologous tumor extract. This factor was found in 18 patients, who were equally distributed between the two therapeutic groups.In the group with synergistic factor, the percentage with lymph node involvement appeared greater (83% compared with 68% among those patients who had no SS factor), and the incidence of distant metastases was also raised (44% compared with 21%). This factor seemed to indicate a poor prognosis. Moreover, there was a difference in the results between the two therapeutic groups in patients with synergistic factor. Of nine patients undergoing conventional treatment, six developed metastases, whereas only two of the nine patients also receiving poly A · poly U developed metastases. The same trend was observed throughout the trial population.     

Tumour-associated antigens in culture medium of malignant melanoma cell strains

Summary The presence of melanoma-associated antigens naturally shed from cultured melanoma cells in spent culture medium was investigated by means of a leukocyte migration test and culture medium from four melanoma and two control cell strains.Leukocytes from 29/64 melanoma patients showed a positive reaction with spent culture medium from at least one melanoma cell strain, whereas leukocytes from only 4/25 patients with other cancers and 1/30 normal donors reacted. On the other hand, leukocytes from only 8/51 melanoma patients reacted with control culture medium. Only melanoma patients' leukocytes reacted with two or more of the melanoma cell strains used. Culture media from two melanoma cell strains were more reactive (25.3% and 29.4% positive tests with melanoma patients' leukocytes) than others (12.5% and 17.2% positive tests); this may represent either a qualitative difference (i.e., different antigens) or a quantitative one (i.e., different levels of antigen expression according to tissue culture conditions). Both inhibition and stimulation of migration were observed, but with one exception, on a given occasion, leukocytes from the same donor always reacted in the same way (i.e., either inhibition or stimulation). Migration stimulation was observed mainly with melanoma patients' leukocytes, and more especially when leukocytes were sampled from patients within a few weeks from tumour removal; migration stimulation may thus reflect a particular state of sensitization in patients.From the evidence obtained in these studies, it is concluded that spent culture medium from melanoma cell strains contains melanoma-associated antigen (s) that is (are) reactive in the leukocyte migration test and that this may contribute to the study of specific antitumour reactivity in patients and to the study and purification of tumour-associated antigens by providing an homogeneous source of antigens spontaneously released from tumour cells in conditions close to natural ones.     

Silencing of the MEG3 gene promoted anti-cancer activity and drug sensitivity in glioma

Aberrant expression of MEG3 has been shown in various cancers. The purpose of this study is to evaluate the effect of MEG3 on glioma cells and the use of potential chemotherapeutics in glioma by modulating MEG3 expression. Cell viability, migration and chemosensitivity were assayed. Cell death was evaluated in MEG3 overexpressing and MEG3 suppressed cells. MEG3 expression was compared in patient-derived glioma cells concerning IDH1 mutation and WHO grades. Silencing of MEG3 inhibited cell proliferation and reduced cell migration while overexpression of MEG3 promoted proliferation in glioma cells. MEG3 inhibition improved the chemosensitivity of glioma cells to 5-fluorouracil (5FU) but not to navitoclax. On the other hand, there is no significant effect of MEG3 expression on temozolamide (TMZ) treatment which is a standard chemotherapeutic agent in glioma. Suppression of the MEG3 gene in patient-derived oligodendroglioma cells also showed the same effect whereas glioblastoma cell proliferation and chemosensitivity were not affected by MEG3 inhibition. Further, as a possible cell death mechanism of action apoptosis was investigated. Although MEG3 is a widely known tumour suppressor gene and its loss is associated with several cancer types, here we reported that MEG3 inhibition can be used for improving the efficiency of known chemotherapeutic drug sensitivity. We propose that the level of MEG3 should be evaluated in the treatment of different glioma subtypes that are resistant to effective drugs to increase the potential effective drug applications.     

Colorectal cancer diagnosis by a direct leukocyte migration test using a panel of tumour extracts

Summary In a direct leukocyte migration test, peripheral blood leukocytes were pulsed with a high dose (2.5 and 0.5 mg/ml) of 3 M KCl extracts from 5 different colorectal tumours as well as with one 3 M KCl extract of normal colonic mucosa. Patients showing a pathological migration index (0.80 and 1.17), with 3 or more out of 5 tumour extracts, were considered as positives.With this test mode 93% (55/59) of patients with colorectal carcinomas were reactive, irrespective of the tumour stage, while only 7% (2/27) of patients with non-malignant colorectal diseases showed a positive reaction. Patients with malignant and non-malignant diseases of other organs were reactive in 2–3% of cases. No positive reactivity was observed with leukocytes from 37 healthy volunteers. Pulsing leukocytes with the normal colonic mucosal extract, a pathological migration index was found in about 20% of colorectal cancer patients, but not in healthy volunteers.Evaluating 10 single tumour extracts individually, reactivity of cancer patients' leukocytes ranged from 65–89% of tests, the difference being not statistically significant. Leukocytes from healthy volunteers showed a pathological migration index with the different extracts in 0–6% of tests.With the leukocyte migration test we could not differentiate between tumours of the colon, sigma or rectum. Patients bearing tumours in any part of the large bowel showed pathological leukocyte migration with extracts of colon-, sigma- and rectum tumours. When the cross-reactivity study was extended to tumours of the gastrointestinal tract, it was found that patients with colorectal tumours were reactive, in a high percentage of tests, with extracts of gastric tumours, but gastric as well as oesophageal and pancreatic cancer patients' leukocytes only reacted occasionally with colorectal tumour extracts.In the follow-up study, a positive reactivity was still found 10–14 days after surgery in 27/31 patients. After more than 2 months, the frequency of positive reactivity decreased to 10/70 cases. Patients with local recurrence or metastases exhibited positive reactivity in 6/7 cases.Abbreviations LMT leukocyte migration test - LM leukocyte migration - LMI leukocyte migration inhibition - LME leukocyte migration enhancement - MI migration index     

Changes induced with varying solutions of normal plasma on granulocyte capillary tube migration

In the present work we have evaluated the effect of decreasing concentrations of normal plasma in TC 199 on leukocyte capillary migration. From the 1st to the 4th hour of migration, normal plasma at 100%-80%-60%-40% concentration, significantly inhibits leukocyte migration. The inhibitory effect is lost, only by a few plasmas, at 60% and 40% concentration, during the following migration hours. Normal plasma at 20% and 10% concentration doesn't show any inhibitory effect on leukocyte capillary migration. Therefore, for clinical studies on pathological plasmas we recommend the use o plasma at 20% or 10% concentration to avoid aspecific inhibition.     

Postoperative Depression of Tumour-directed Cell-mediated Immunity in Patients with Malignant Disease

Leucocytes from 46 melanoma patients, 45 breast carcinoma patients, and 95 control donors were tested by the leucocyte migration test against the supernatants of homogenates of malignant melanomas, breast carcinomas, simple breast tumours, and breasts showing simple cystic disease. By comparison with controls inhibition of migration occurred significantly more frequently when tumour patients'' leucocytes were exposed to extracts of histogenetically similar tumours.Cell-mediated immunity to tumour-associated antigens was measured in 12 patients with breast carcinoma and 12 with malignant melanoma immediately before surgical operation and in the postoperative period. All patients tested before operation showed significant inhibition of migration on contact with extracts of histogenetically similar tumours. Postoperatively the degree of leucocyte migration inhibition was reduced in all patients with melanoma and breast carcinoma. Significant inhibition of leucocyte migration returned in most patients 6-22 days after operation.     

Restoration of antineoplastic reactivity of the leukocytes after preincubation]

Specific antitumor sensitization in patients with carcinoma of the stomach was revealed by means of the leukocyte migration inhibition test. After 24-hour preincubation at 4 degrees C in a serum-free medium the areactive leukocytes from patients with carcinoma of the stomach (stages III--IV) acquired the ability to react specifically to allogeneic antigens of the tumor of the same localization. Preincubation did not influence the inhibition of migration of leukocytes in non-tumor patients by stomach carcinoma antigens. The supernatant of preincubated leukocytes of tumor patients contained substances that inhibited migration of indicator leukocytes.     

Tumour suppressor genes in chemotherapeutic drug response

Since cancer is one of the leading causes of death worldwide, there is an urgent need to find better treatments. Currently, the use of chemotherapeutics remains the predominant option for cancer therapy. However, one of the major obstacles for successful cancer therapy using these chemotherapeutics is that patients often do not respond or eventually develop resistance after initial treatment. Therefore identification of genes involved in chemotherapeutic response is critical for predicting tumour response and treating drug-resistant cancer patients. A group of genes commonly lost or inactivated are tumour suppressor genes, which can promote the initiation and progression of cancer through regulation of various biological processes such as cell proliferation, cell death and cell migration/invasion. Recently, mounting evidence suggests that these tumour suppressor genes also play a very important role in the response of cancers to a variety of chemotherapeutic drugs. In the present review, we will provide a comprehensive overview on how major tumour suppressor genes [Rb (retinoblastoma), p53 family, cyclin-dependent kinase inhibitors, BRCA1 (breast-cancer susceptibility gene 1), PTEN (phosphatase and tensin homologue deleted on chromosome 10), Hippo pathway, etc.] are involved in chemotherapeutic drug response and discuss their applications in predicting the clinical outcome of chemotherapy for cancer patients. We also propose that tumour suppressor genes are critical chemotherapeutic targets for the successful treatment of drug-resistant cancer patients in future applications.     

Development of specific cellular immunoreactivity in typhoid fever.

Development of cellular immunoreactivity to Salmonella typhi and Salmonella paratyphi-A was studied by the leukocyte migration inhibition test in 9 patients with typhoid fever and in 2 patients with paratyphoid fever. Cellular reactivity could be demonstrated from the first days of the disease in all the subjects. The most pronounced migration inhibition was observed during the febrile period. It is suggested that specific cellular reactivity may play a pathogenetic role in typhoid fever.     

Role of pure and biologically active amoebal RNA in assessment of lymphokines: a report of 55 ALA cases.

Amoebic liver abscess cases (55) were assessed for release of lymphokines (LMIF) using pure and biologically active amoebal RNA of axenic Entamoeba histolytica (NIH: 200) obtained with cesium chloride centrifugation. Lymphokines released by T lymphocytes in response to both amoebal RNA and whole amoebic lysate (WAL) were tested by leukocyte migration inhibition test (LMIT) on blood samples from amoebic liver abscess cases. A significant increase was observed in the release of lymphokine and 100% positivity was observed with amoebal RNA compared to whole amoebic extract with a positivity of only 78%. The difference between means leukocyte migration inhibition of the above two with regards to release of lymphokine was highly significant (P less than 0.001). This shows that patients had high degree of leukocyte sensitization to amoebal RNA of E. histolytica compared to whole amoebic lysate. These findings suggest that the amoebal RNA plays an important role as a potent antigen in the elicitation of cell mediated immune responses in amoebic liver abscess cases.     

Mahanine synergistically enhances cytotoxicity of 5-fluorouracil through ROS-mediated activation of PTEN and p53/p73 in colon carcinoma

5-Fluorouracil (5-FU) alone or in combination with other drugs is the main basis of chemotherapeutic treatment in colorectal cancer although patients with microsatellite instability generally show resistance to 5-FU treatment. The present investigation is focussed on the mechanistic insight of a pure herbal carbazole alkaloid, mahanine, as a single or in combination with 5-FU in colon cancer. We demonstrated that mahanine-induced apoptosis involved reactive oxygen species (ROS)-mediated nuclear accumulation of PTEN and its interaction with p53/p73. Mahanine and 5-FU in combination exerted synergistic inhibitory effect on cell viability. This combination also enhanced ROS production, increased tumour suppressor proteins and suppressed chemo-migration. Taken together, our results revealed that mahanine can be a potential chemotherapeutic agent with efficacy to reduce the concentration of toxic 5-FU in colon cancer.     

Ability of renal carcinoma tissue extract to induce leukocyte migration inhibition in patients with nonmetastatic renal carcinoma: Correlation with clinical and histopathological findings

Summary Soluble extracts prepared from 24 renal carcinomas were tested for their ability to induce inhibition of migration of leukocytes from 27 patients with nonmetastatic hypernephroma using the capillary tube leukocyte migration technique. No correlations were found between the histopathology of the carcinomas tested and this ability. However, highly significant correlations were demonstrated between the existence of nonmetastatic tumor and tumor-directed, cell-mediated hypersensitivity in the tumor donors and the ability of tumor tissue extracts from such patients to induce leukocyte migration inhibition in allogeneic tests.The capacity of renal carcinoma tissue extracts to induce leukocyte migration inhibition in renal carcinoma patients seems in allogeneic combinations to be an in vitro correlate of in vivo tumor antigenicity.     

Cellular hypersensitivity to human pancreatic B-cell clone in diabetes mellitus and its relationship to the presence of islet cell antibodies

A leukocyte migration inhibition test on the human pancreatic B-cell clone (JHPI-1) was performed in 13 IDDM patients with islet cell cytoplasmic antibody (ICCA) and/or islet cell surface antibody (ICSA), 15 IDDM patients without ICCA or ICSA, 34 NIDDM patients and 17 healthy controls. The mean values for the migration index (M.I. %) in each group were 85.4 +/- 6.9, 89.1 +/- 10.9, 98.3 +/- 7.9 and 100.0 +/- 8.5. The M.I. values were significantly decreased in IDDM patients than in NIDDM patients and controls irrespective of whether or not there were islet cell antibodies in the patients' sera. When M.I. values less than 0.83 (Mean-2 S.D.) were taken as indicative of inhibition, the percentage of IDDM and NIDDM patients with migration inhibition were 32% and 0% respectively. And the decreased M.I. values in IDDM patients proved not to be due to non-specific migration inhibition by normal M.I. values, with the human fetal lung fibroblast cells (W 138) as antigen. Our data suggested that the lymphocytes of IDDM patients might be sensitized by pancreatic B-cell antigen(s) present in the JHPI-1 cells, which promoted leukocyte migration inhibition. No correlation between the migration indices and duration of diabetes mellitus in IDDM patients was observed (r = 0.254, Y = 84.9 + 0.49 X). LMT to JHPI-1 seems to be useful in detecting the abnormal cell-mediated immunity even in patients with longstanding IDDM.     

Detergent dissection of membrane proteins of Entamoeba histolytica and its effect on lymphokine release in in vitro.

Fifty-two amoebic liver abscess cases were assessed for the release of lymphokines (LMIF) using detergent dissected membrane proteins (DDMP) of axenic Entamoeba histolytica (NIH:200) obtained with sodium deoxycholate treatment. Lymphokines release by T lymphocytes in response to both DDMP and whole amoebic lysate (WAL) was tested by leukocyte migration inhibition test on blood samples from amoebic liver abscess cases. A significant increase was noted in the release of LMIF and 100% positivity was observed with DDMP compared to whole amoebic extract with a positivity of 73%. The difference between means of the above two with regards to release of LMIF was found to be highly significant (P less than 0.005). This shows the patients had high degree of leukocyte sensitization to surface antigens of E. histolytica compared to the whole amoebic lysate. These findings suggest that the antigens shed might have important role as a potent antigen in elicitation of CMI response in amoebic liver abscess cases.     

Colony stimulating factor levels in human serum and urine following chemotherapy.

The colony stimulating activity (CSA) of serum and urine specimens from 10 patients with lymphomas before and after treatment with standard multidrug chemotherapeutic regimens has been studied. Prominent rises in the CSA in the urine of eight of these 10 patients correlating temporally with initiation of therapy was demonstrated when compared to pretreatment control values. Hourly serum specimens were collected following the administration of intravenous chemotherapeutic agents in two of these patients. In one of these two patients there was a prominent and early rise in CSA which returned to control values by the 12th hr after intravenous drug administration. In two patients who developed little or no increase in CSA during therapy, urine specimens were studied for the presence of inhibitors to colony stimulating factor. Aliquots of urine from each of these two patients when mixed with a standard control urine of known CSA, produced marked inhibition.