Increased chemosensitivity to doxorubicin of intrinsically multidrug-resistant human colon carcinoma cells by prolonged exposure to verapamil

Resistance modifying agents (RMA) such as verapamil (VER) have proved effective in reversing multidrug resistance (MDR) in many in vitro experimental models, but clinical results with RMA have been disappointing. To clarify this apparent discrepancy we have evaluated the cytotoxic effects of doxorubicin (DOX) plus VER in four human colon carcinoma (HCOC) cell lines (LoVo, DLD-1, SW948, SW1116). These lines were selected on the basis of their levels of mdr1 mRNA being similar to those expressed by HCC obtained from non-drug-treated patients. In all cell lines the sensitising effect of VER on DOX cytotoxicity was schedule-dependent and maximal potentiation of DOX cytotoxicity was obtained by exposure to VER for a time > or = the cells' population doubling time.     

Antileukemic action of buthionine sulfoximine: evidence for an intrinsic death mechanism based on oxidative stress

The glutathione-depleting agent buthionine sulfoximine (BSO) was found to be toxic to some AML blast populations. This toxicity was manifested as the appearance of high levels of reactive oxygen generation in GSH-depleted cells, and later by the loss of mitochondrial membrane potential and an increase in intracellular calcium. Striking heterogeneity in BSO sensitivity was observed in a series of four human AML cell lines, and in fresh leukemic blasts obtained from eight AML patients. In some cases, toxicity was seen at BSO concentrations as low as 1 microM; approximately 100-fold less than the plasma levels achieved in patients treated with BSO as a drug resistance reversing agent. Based on these results we propose that some AML blast populations are unusually dependent on GSH-based antioxidant mechanisms, due to high intrinsic rates of reactive oxygen generation. The mitochondrial respiratory chain is the most likely source of this reactive oxygen. Because toxicity is seen at clinically achievable concentrations of BSO, this agent might have antileukemic activity in patients.     

Effects of buthionine sulfoximine on the outcome of the in utero administration of alcohol on fetal development

The adverse effects of the maternal consumption of alcohol on the fetus have been recognized for centuries. Fetal alcohol syndrome is characterized by pre- and postnatal growth retardation, mental retardation, behavioral deficits, and facial deformities. Despite numerous animal studies, the biochemical mechanism(s) by which alcohol produces teratogenic effects on the developing fetus are not well understood. Several studies have shown that administration of alcohol to adult rats produces a decrease in hepatic levels of glutathione (GSH). In utero administration of alcohol has also been shown to produce a decrease in GSH levels, as well as prenatal growth retardation and intrauterine death. In an effort to determine if GSH may have a vital role in protecting the fetus against the teratogenic effects of alcohol, buthionine (SR)-sulfoximine (BSO) was used to deplete GSH levels in the mother and fetus. Timed pregnant Sprague-Dawley rats were placed on a liquid BioServ diet containing either 0%, 11%, 23%, 29%, 31%, 33%, or 35% ethanol-derived calories, with or without BSO (888 mg/kg/24 hr), starting on day 1 of pregnancy. Another set of mothers were fed lab chow and water as a control group for the liquid diet. The mothers were maintained on the diet until gestation day 21 when they were anesthetized with sodium pentobarbital and the pups delivered by cesarean section. The offspring were counted, weighed, killed, and the brain and liver weighed. The effects of BSO on the alcohol dose-response curves (body weights, brain weights, and litter number) were then determined to ascertain if a depletion in GSH potentiated the effects of alcohol. In utero administration of BSO, aside from the depletion of GSH in the liver and brain in the developing fetus, produced a shift to the left in the alcohol dose-response curve.     

Effect of buthionine sulfoximine, a synthesis inhibitor of the antioxidant glutathione, on the murine nigrostriatal neurons

This study analyzed the effects of acute systemic treatment with buthionine sulfoximine (BSO), a synthesis inhibitor of the antioxidant reduced glutathione (GSH), on dopaminergic neurons of the murine nigrostriatal pathway. Part 1 of the study established a dose-response curve and the temporal pattern of GSH loss and recovery in the substantia nigra and striatum following acute BSO treatment. Part 2 of the study determined the effect of acute BSO treatment on the morphology and biochemistry of nigrostriatal neurons. We found that decreases in GSH levels had profound morphological effects, including decreased catecholamine fluorescence per cell, increased levels of lipid peroxidation and lipofuscin accumulation, and increased numbers of dystrophic axons in dopaminergic neurons of the nigrostriatal pathway. However, no measurable effects were observed in biochemical levels of either dopamine or its metabolites. These changes mimic those that have been reported to occur in the nigrostriatal system of rodents with advancing age. Our data suggest that reduction of GSH via BSO treatment results in the same types of nigrostriatal degenerative effects that occur during the aging process and consequently is a good model system for examining the role of GSH in protecting this area of the brain against the harmful effects of age-related oxidative stress.     

In vivo effects of tumor necrosis factor-alpha or flavone acetic acid in combination with doxorubicin on multidrug-resistant B16 melanoma

BACKGROUND: A soft pancreas with a main pancreatic duct (MPD) with normal diameter has been considered a high risk for pancreatic anastomotic leakage because of a relatively high output of pancreatic juice, but data are lacking. METHODS: An attempt was made to assess the relationship between the consistency of the pancreas, MPD diameter, pancreatic juice output, and pancreatic leakage after partial pancreatoduodenectomy. The pancreatic parenchyma was classified as of soft, intermediate, and hard consistency in 70 consecutive patients undergoing operation (groups 1, 2, and 3, respectively) by one surgeon. The MPD diameter was determined by means of endoscopic pancreatography or abdominal ultrasonography. Pancreatic juice output was measured for 21 days after operation by using a catheter inserted into the MPD. Anastomotic leakage was identified radiologically by using contrast medium. RESULTS: The mean (SD) pancreatic juice output during a period of 10 days (postoperative days 5 to 14) was 1554 (1073) ml in group 1 (n = 29), 1513 (1060) ml in group 2 (n = 13), and 187 (220)ml in group 3 (n = 28) (groups 1 and 2 versus group 3, p < 0.0001). The MPD diameter was 3.0 (1.6) mm in group 1, 5.9 (2.5) mm in group 2, and 6.6 (2.6) mm in group 3 (group 1 versus groups 2 and 3, p = 0.0001). Anastomotic leaks occurred in four (14%) patients in group 1, three (23%) in group 2, and none in group 3 (p < 0.05). CONCLUSIONS: Patients with a pancreatic parenchyma with an intermediate or normal consistency produced more pancreatic juice and had a higher leak rate.     

Taxol-enhanced cytotoxic effect of radiation in human promyelocytic leukemia cells: relative resistance of multidrug-resistant HL-60 cells in vitro

Cytotoxic effects of sequential taxol (paclitaxel) and X-irradiation on drug-sensitive human cultured promyelocytic leukemia (HL-60) cell line and its multidrug-resistant sublines were examined using photometric MTT test and flow cytometry. Paclitaxel (at concentrations 1-10 nmol) stimulated the cytotoxic effect of irradiation in HL-60 and to a lesser extent also in HL-60/ADR, but not in HL-60/VCR cells. The stimulation of radiation-induced cytotoxic effect by paclitaxel correlated with its potential to induce cell cycle and viability alterations identified with flow cytometric analysis (i.e. increased propidium iodide staining, increased side scatter, decreased forward angle scatter, accumulation of necrotic cell detritus, apoptotic pre-G0 cells and cells in the G2/M phase of the cell cycle).     

Comparison of the cytotoxic effects of corticoids on the neoplastic B cells

Many clinical trials have been engaged to prove the benefits of new drugs in the treatment of hematological tumours. However, no real progress have occurred in diseases such as multiple myeloma, the association of melphalan and prednisone is still the mainstay of the treatment. During all these years, the family of glucocorticoids have not been totally studied. Their efficiency in the cure of lymphoid malignancies has been early recognised, but still to be based on their anti-inflammatory potency for the dosages. Only few works reported the comparison between members of this family. We demonstrate in this work, in vitro, with a cell line of medium sensibility and a B cell of tumoral origin grew up in our laboratory, that exists some differences in the anti-neoplastic potency of the more commonly used corticoids. If the order in which we can class these drugs is not surprising and empirically known, the importance of the differences observed need a special attention. We also found that these drugs might have stimulatory effects, at various degree in function of their concentrations, on the proliferation of the B cell lines. Theses side effects coupled to the efficiency variations of each corticoid present the need of paying more attention to the choice of the molecule implied in the chemotherapy.     

Detection of cytotoxic activity on Vero cells in clinical isolates of Serratia marcescens

Cytotoxin production was studied in 60 Serratia marcescens strains isolated from hospitalized patients. Association of cytotoxic activity with serotype, source of isolation and presence of plasmids was also evaluated. Thirteen of the 60 S. marcescens strains produced a cytotoxic effect on Vero cells. These strains were isolated from distinct clinical sources and classified into seven different serotypes (O1:H7; O4:NM; O10:NT; O19:NM; O6,14:H4; O6,14:NM and O6,14:H1). No relationship was observed between cytotoxic activity and clinical source or serotypes of the strains. Plasmids from five cytotoxin-producing S. marcescens strains were transferred to E. coli K12/711. The transconjugants did not exhibit cytotoxicity, indicating that the cytotoxic effect is not plasmid-mediated among these strains. Although a cytotoxic activity was demonstrated in filtrates of some S. marcescens strains, further studies should be performed to assess the role of this toxin in pathogenesis.     

N-acetylcysteine improves cytotoxic activity of cirrhotic rat liver-associated mononuclear cells

Liver cirrhosis, which is associated with decreased plasma and hepatic glutathione (GSH), has been reported to cause the suppression of NK activity in peripheral blood mononuclear cells. Since low GSH levels in lymphocytes are known to alter lymphocyte function, we examined the correlation between intracellular GSH levels and the cytotoxic activity of liver-associated mononuclear cells (liver MNC). We show here that rat liver contains a highly active population of NK cells (CD3- NKR-P1 + cells) that kill Yac-1 in vitro and that the cytotoxic activity of this NK population is directly proportional to liver MNC GSH. This proportionality is independent of the methods used to alter GSH level. Thus, in vitro treatment of liver MNC with buthionine sulfoximine to lower GSH levels lowers the cytotoxic activity. MNC from cirrhotic liver, in which implanted tumor cells grow faster, have both low GSH levels and low cytotoxicity, and supplementation of cirrhotic liver MNC with N-acetylcysteine raises GSH levels and increases cytotoxicity. These findings suggest a physiologic mechanism, i.e. decreased GSH, may be causally associated with the increased incidence of hepatoma in cirrhotic individuals and the increased growth of hepatoma cells in cirrhotic animals. Thus, we suggest that the GSH is important to the optimal functioning of the hepatic immunity that protects against hepatoma development.     

Dexamethasone-induced cytotoxic activity and drug resistance effects in androgen-independent prostate tumor PC-3 cells are mediated by lipocortin 1

We studied the seated buttock pressure distribution in six paraplegic patients by means of computerized pressure mapping. They were all male and their age ranged from 18 to 48 years old. Their level of paralysis varied from Th5 to L1. Five kinds of wheelchair cushions were studied: an air cushion, a contour cushion, a polyurethane foam cushion, a Cubicushion (which is made of polyurethane foams) and a silicone gel cushion. A tactile sensor consisting of 2064 matrices was used for measuring the buttock pressure distribution and the data was analyzed on a personal computer. Peak pressures measured for each cushion were as follows (in descending order): the Cubicushion, the polyurethane foam cushion, the contour cushion, the silicone gel cushion, and the air cushion. The areas of total contact measured for each cushion were as follows (in descending order): the air cushion, the silicone gel cushion, the polyurethane foam cushion, the contour cushion and the Cubicushion. Based on these findings, we conclude that the most advantageous cushion is the air cushion or the silicone gel cushion. Likewise, we conclude that the Cubicushion is not practical for pressure sore prevention.     

Detection of cytotoxic effects of Clostridium novyi type A on bovine kidney cells

The cytotoxic effects of a toxic preparation from Clostridium novyi type A were demonstrated on tissue-cultured bovine kidney cells. The cytotoxic response was dose-dependent and could be neutralized by homologous antiserum. Scanning electron microscopy revealed damaged kidney cell surfaces. These findings indicated that the cytotoxicity may contribute to the formation of the foci in bovine tissue during an infection with C. novyi.     

Direct cytotoxic and cytoprotective effects of some drugs on isolated cells of gastric mucosa from rats

Toxic effect of ethanol and indomethacin, and the cytoprotective effect of prostacyclin were studied. In absence of other aggressive factors, the indomethacin was found to be non-toxic, but it could enhance the necrotizing effect of ethanol, this effect could be counteracted by prostacyclin preincubation. These observations suggest that there is an endogenous prostaglandin system in rat gastric mucosal cells that may protect against the toxic effect of indomethacin; administered prostacyclin may have a cytoprotective effect when this endogenous prostaglandin system's protective function does not more function. Gastric mucosal cytoprotection may include cellular level processes, which are able to operate without the existence of tissue integrity.     

Inhibition of fos-jun-DNA complex formation by dihydroguaiaretic acid and in vitro cytotoxic effects on cancer cells

The main cause of acquired inguinal hernia is weakness of Fruchaud's deep muscolofascial floor, following metabolically-determined collagen disorders. A technique for the anterior reinforcement of this structure with polypropylene mesh is described here. Following intermuscular decollement, the mesh is placed in direct contact with the surface formed by the transversalis fascia and the transversus abdominis muscle and stretched as extensively as possible. Because the posterior aspect of the inguinal canal is the true barrier to abdominal pressure, the author believe that its direct reinforcement, without interposition of the internal oblique muscle, constitutes the most correct anatomo-surgical approach to hernia repair. This is the case for both indirect hernias, in which the internal ring is reconstructed at a deeper level, and for direct hernias, in which the "tent effect" of the prosthesis is prevented. Ninety-two primary inguinal hernias (56 indirect, 29 direct and 7 direct and indirect) in 87 patients were repaired with this technique. Seventy-nine patients were followed up from 2 to 24 months. Early complications included: 7 ecchymosis, 3 seromas, 2 subcutaneous infections, 3 testicular swellings. Incision and testicular pain for longer than 6 months occurred in 2 cases. No prosthetic infections or recurrences have been detected up to the present.     

Effect of enkephalins and endorphins on cytotoxic activity of natural killer cells and macrophages/monocytes in mice

The effect of [Met5]enkephalin, [Leu5]enkephalin, proenkephalin, dynorphin-(1-17) or beta-endorphin on the cytotoxic (51Cr release assay) activity of natural killer cells and macrophages/monocytes was studied in mice. It was found that a single i.p. injection of [Met5]enkephalin, [Leu5]enkephalin, beta-endorphin, dynorphin or proenkephalin as well as repeated treatment with both enkephalins increased natural killer cell activity. In vitro only [Met5]enkephalin stimulated natural killer cells. Opioid peptides did not affect the cytotoxic activity of macrophages/monocytes. In addition to functional alterations, both enkephalins and beta-endorphin increased the percentage of cells with natural killer phenotype. The results of this study suggest that the increase in natural killer cytotoxicity after opioid peptides injected once or for 14 days may result from an increased number of natural killer cells in the spleen.     

Saccharide-assisted delivery of cytotoxic liposomes to human malignant cells

The overexpression of lectins by malignant cells was applied for in vitro targeting of liposomes equipped with a saccharide vector and loaded in the lipid phase with a lipid derivative of anticancer agent sarcolysine. The lectin specificity of human leukemia HL-60 and human lung adenocarcinoma ACL cells was revealed by tests with fluorescein-labeled sugar probes. With the help of fluorescent lipid dye it was shown that active saccharide ligands increased the level of the vectored liposome binding to malignant cells by 50-80% as compared to liposomes without vector or with inactive one. The degree of liposome/cell membrane fusion was monitored fluorometrically and was shown to be complete and independent of the vectors. The targeted drug-loaded liposomes had the cytotoxic activity 2-4 times higher as compared to the vector-free ones.     

Establishment by adriamycin exposure of multidrug-resistant rat ascites hepatoma AH130 cells showing low DT-diaphorase activity and high cross resistance to mitomycins

A resistant subline (AH130/5A) selected from rat hepatoma AH130 cells after exposure to adriamycin (ADM) showed remarkable resistance to multiple antitumor drugs, including mitomycin C (MMC) and porfiromycin (PFM). PFM, vinblastine (VLB), and ADM accumulated in AH130/5A far less than in the parent AH130 (AH130/P) cells. AH130/5A cells showed overexpression of P-glycoprotein (PGP), an increase in glutathione S-transferase activity, and a decrease in DT-diaphorase and glutathione peroxidase activity. The resistance to MMC and VLB of AH130/5A cells was partly reversed by H-87, an inhibitor of PGP. Buthionine sulfoximine, an inhibitor of glutathione synthase, did not affect the action of MMC. tert-Butylhydroquinone induced DT-diaphorase activity, increased PFM uptake, and enhanced the growth-inhibitory action of MMC in AH130/5A cells. Dicumarol, an inhibitor of DT-diaphorase, decreased PFM uptake and reduced the growth-inhibitory action of MMC in AH130/P cells. These results indicated that the adriamycin treatment of hepatoma cells caused multifactorial multidrug resistance involving a decrease in DT-diaphorase activity.     

Role of hydrogen peroxide in the cytotoxic effects of UVA/B radiation on mammalian cells

Inhibitory effect of four carotenoids prevalent in human blood and tissues against the formation of colonic aberrant crypt foci was examined in Sprague-Dawley rats. They received three intrarectal doses of N-methylnitrosourea in weak 1, and a daily gavage of de-escalated doses of carotenoids during weeks 2 and 5. Lycopene, lutein, alpha-carotene and palm carotenes (a mixture of alpha-carotene, beta-carotene and lycopene) inhibited the development of aberrant crypt foci quantitated at week 6, but beta-carotene did not. The results suggested that lycopene and lutein in small doses may potentially prevent colon carcinogenesis.     

Diisopropylglutathione ester protects A549 cells from the cytotoxic effects of sulphur mustard

The A549 cell line was used to assess the ability of diisopropylglutathione (DIPE) to protect against a 100 microM challenge dose of sulphur mustard (HD) using gentian violet (GV), thiazolyl blue (MTT) and neutral red (NR) assays as indicators of cell culture viability. As part of a continuing study of the efficacy of protective nucleophiles as candidate treatments for HD poisoning, several different combinations of protectant and HD were used to determine the optimal means of protecting A549 cells from the effects of HD. It was found that DIPE (4 mM) could protect cells against the effects of HD though for optimal effect, DIPE had to be present at the time of HD challenge. Cultures protected with DIPE were up to 2.9-fold more viable than HD exposed cells 48 h after HD challenge when using the GV, MTT and NR assays to assess viability. Observations by phase contrast microscopy of GV stained cultures confirmed these findings. Pretreating A549 cultures with DIPE for 1 h followed by its removal prior to HD challenge did maintain cell viability, though at a relatively low level (only up to 1.4-fold more viable than HD only exposed cells). DIPE was also able to protect HD exposed A549 cultures when added to cell cultures at intervals of up to 12 to 15 min after the initial HD exposure, though viability tended to decrease over this period, so that at 1 h, addition of DIPE did not maintain the viability of the cultures. This is the first such report of the anti-HD protectant properties of DIPE in A549 cells. It is concluded that the protection observed against HD is probably largely due to extracellular inactivation of HD by DIPE.