Studies on the mechanisms of invasion in cancer. I. Isolation and purification of a factor chemotactic for cancer cells

A substance chemotactic for cancer cells was extracted from the pseudoglobulin fraction of some tumor tissues of animal and human origin. It was highly purified by column chromatography using Sephadex G-50 and CM-Sephadex and then by disc electrophoresis; it behaved as a homogenous substance on disc electrophoresis. The substance was a protein with a molecular weight of approximately 70,000. It was thermolabile and had no proteolytic activity. The material was similarly active for rat ascites hepatoma AH109A cells, mouse ascites hepatoma MH134 cells and mouse myeloid leukemia C-1498 cells, suggesting a common chemotactic action. It was ineffective for polymorphonuclear leukocytes of rats. No such chemotactic activity was demonstrated by the protein fraction from normal skin and muscle.     

Effect of adriamycin on the metabolism of heart slices

Incubation of Adriamycin (ADR) with either microsomal fractions or human erythrocytes results in the generation of reactive oxygen compounds (ROC). In mice administration of ADR causes lipid peroxidation and soluble sulfhydryl depletion of cardiac tissue. This study determined the effect of ADR on the metabolism of rat heart slices in vitro. Resting heart slices had a high rate of Krebs cycle activity as indicated by the oxidation of ¹⁴C-6-glucose and a low rate of hexose monophosphate shunt activity (HMPS) as indicated by the oxidation of ¹⁴C-1-glucose compared to ¹⁴C-6-glucose. Adriamycin (100μg/ml) stimulated HMPS activity 2 fold but did not change Krebs cycle activity. The HMPS pathway of heart slices incubated under anaerobic conditions was not stimulated by ADR. Unexpectedly, heart slices incubated with ¹⁴C-formate and catalase had a high baseline oxidation of this substrate which was not augmented by ADR. These studies indicate that ADR stimulates the HMPS activity of rat heart without altering Krebs cycle activity. The observation that the stimulation is dependent on oxygen suggests the generation of ROC. Resting heart slices have a high baseline production of H₂O₂ which does not appear to be associated with stimulation of the HMPS pathway. This observation suggests that H₂O₂ is not degraded adequately by the HMPS pathway in cardiac tissue. These studies are consistent with the concept that ROC may be important in the cardiac toxicity of ADR related in part to a sensitivity of cardiac tissue to oxidant injury.     

Carbohydrate metabolism in the kidneys of rats with dimethylnitrosamine-induced tumors]

Under study was anaerobic glycolysis, the activity of hexokinase, lactate dehydrogenase, gluco-6-phosphatase in rat kidneys in dimethylnitrosamine (DMNA) induced tumors in them. DMNA was administered perorally in the dosage of 10 mg/Kg during 4 weeks (25 mg per rat). Following 8 months tumors developed in the renal cortical substance. The tumor tissue, the renal cortical substance adjacent to the tumor, and the renal cortical substance without macroscopic tumor signs have been studied. An increased glycolysis, hexokinase and lactatedehydrogenase activity were noted, whereas the activity of glucoso-6-phosphatase was diminished in all tissues under examination. Changes of all indices were mostly pronounced in tumor tissue and least significant--in the renal cortical layer without any macroscopic tumor signs.     

Autocrine tumor growth regulation and tumor-associated hypoglycemia in murine melanoma B16 in vivo

A substance immunochemically cross-reactive with insulin (SICRI) appears in melanoma B16 growing in diabetic and nondiabetic C57BL/6 mice. Progression of tumor size is paralleled by the increase of SICRI levels in the serum of both diabetic and nondiabetic animals; this increase correlates with a decreased concentration of circulating glucose and an elevated concentration of growth hormone in blood. Melanoma B16 grown under serum-free culture conditions secretes SICRI into the medium. Affinity-purified SICRI stimulates glucose uptake by rat epididymal adipocytes and competes with radiolabeled insulin for binding to these cells. Low concentrations of SICRI enhance growth of cultured melanoma B16 cells, whereas high concentrations of this substance have inhibitory growth effects on these cells. Porcine insulin, human insulin-like growth factors I and II, human growth hormone, platelet-derived growth factor, epidermal growth factor, and fibroblast growth factor have negligible influence on growth of melanoma B16.     

Effect of a tumour-derived lipid-mobilising factor on glucose and lipid metabolism in vivo

Treatment of ex-breeder male NMRI mice with lipid mobilising factor isolated from the urine of cachectic cancer patients, caused a significant increase in glucose oxidation to CO2 compared with control mice receiving phosphate buffered saline. Glucose utilisation by various tissues was determined by the 2-deoxyglucose tracer technique and shown to be elevated in brain, heart, brown adipose tissue and gastrocnemius muscle. The tissue glucose metabolic rate was increased almost three-fold in brain, accounting for the ability of lipid mobilising factor to decrease blood glucose levels. Lipid mobilising factor also increased overall lipid oxidation, as determined by the production of 14CO2 from [14C carboxy] triolein, being 67% greater than phosphate buffered saline controls over a 24 h period. There was a significant increase in [14C] lipid accumulation in plasma, liver and white and brown adipose tissue after administration of lipid mobilising factor. These results suggest that changes in carbohydrate metabolism and loss of adipose tissue, together with an increased whole body fatty acid oxidation in cachectic cancer patients, may arise from tumour production of lipid mobilising factor.     

Anti-interleukin-6 receptor antibody prevents muscle atrophy in colon-26 adenocarcinoma-bearing mice with modulation of lysosomal and ATP-ubiquitin-dependent proteolytic pathways

Progression of skeletal muscle atrophy is one of the characteristic features in cancer patients. Interleukin-6 (IL-6) has been reported to be responsible for the loss of lean body mass during cancer cachexia in colon-26 adenocarcinoma (C-26)-bearing mice. This study was carried out to elucidate the intracellular proteolytic pathways operating in skeletal muscle in C-26-bearing mice, and to examine the effect of anti IL-6 receptor antibody on muscle atrophy. On day 17 after tumor inoculation, the gastrocnemius muscle weight of C-26-bearing mice had significantly decreased to 69% of that of the pair-fed control mice. This weight loss occurred in association with increases in the mRNA levels of cathepsins B and L, poly-ubiquitin (Ub) and the subunits of proteasomes in the muscles. Furthermore, enzymatic activity of cathepsin B+L in the muscles also increased to 119% of the control. The administration of antimurine IL-6 receptor antibody to C-26-bearing mice reduced the weight loss of the gastrocnemius muscles to 84% of that of the control mice, whose enzymatic activity of cathepsin B+L and mRNA levels of cathepsin L and poly-Ub were significantly suppressed compared with those of the C-26-bearing mice. Our data indicate that both the lysosomal cathepsin pathway and the ATP-dependent proteolytic pathway might be involved in the muscle atrophy of C-26-bearing mice. The results also suggest that anti IL-6 receptor antibody could be a potential therapeutic agent against muscle atrophy in cancer cachexia by inhibiting these proteolytic systems. © 1996 Wiley-Liss, Inc.     

Autoradiographic imaging of rat sarcoma in different anatomical sites using 2-[14C]deoxyglucose

As sarcomas are known to have accelerated glycolysis, we used the radiolabeled glucose analogue 2-deoxy-d-[U-14C]glucose in autoradiographic imaging studies of a methylcholanthrene-induced rat fibrosarcoma placed in an i.m. site, and in models of pulmonary and hepatic metastases. Fifty muCi of 2-deoxy-d-[U-14C]glucose were injected i.p. into groups of rats bearing tumors in these three sites; sacrifice of animals for imaging was carried out 45 min later. Excellent imaging of sarcoma tissue in all three anatomical sites was obtained, with high visual contrast compared to the normal tissue background. Using densitometry of autoradiographs, tumor/tissue ratios were 7.1 for i.m. tumors, 3.8 for pulmonary metastases, and 2.8 for hepatic metastases. Autoradiographic imaging of sarcomas may be obtained based upon avidity of neoplastic tissue for the glucose analogue 2-deoxy-d-[U-14C] glucose. Such imaging is not dependent upon anatomical site and reproducibly images rat sarcomas in muscle, lung, and liver.     

Purification of a cytotoxic factor from human and rat tissues.

Human and rat normal tissues and tumours have been studied for the presence of toxic substances, possibly of importance in the development of cachexia in patients with cancer and other chronic diseases. The toxic effect of tissue extracts was gauged by measuring the inhibition of growth of mouse L-cells in 1-ml cultures, as revealed by reduced incorporation of [14C]leucine into cell protein. A common cytotoxic substance of mol. wt. approximately 700 daltons was isolated from all rat and human tissues tested, including tumours. The isolation procedure involved tissue homogenization, followed by pressure dialysis, gel filtration of concentrated pressure dialysates, cation exchange chromatography, and thin layer chromatography. Amounts isolated from different tissues varied by a factor of 3. The purified substance reacted with ninhydrin and a few other reagents for amino groups. It was completely resistant to acid and enzymatic hydrolysis. The evidence thus suggests that the substance is an amine. It is toxic to L-cells, HeLa cells and normal rat fibroblasts in concentrations of 10-20 muM, producing cell death and lysis during incubation overnight.     

DNA topoisomerase II-mediated interaction of doxorubicin and daunorubicin congeners with DNA

Three groups of doxorubicin and daunorubicin analogues, differing by their substituents on the chromophore and sugar moieties, were used in this study. The 3'-N-unsubstituted (Group 1), 3'-N-acyl (Group 2), and 3'-N-alkyl (Group 3) analogues were tested for: (a) in vivo antitumor activity and in vitro cytotoxicity; (b) cellular or tissue uptake and metabolic conversion; (c) strength of DNA intercalation; and (d) interaction with DNA topoisomerase II (topo-II). Compounds of Group 1 were cytotoxic, were strongly intercalative, and, except for those with C-14 side chain substitution, induced the formation of topo-II-DNA cleavable complexes. As shown previously, esterolysis of C-14-acyl substituents was required to yield a metabolite which can interact with topo-II in the purified system. The C-14-substituted compounds of Group 2 and their C-14-unsubstituted metabolites were cytotoxic. These drugs were weak intercalators, and the C-14-unsubstituted cogeners induced cleavable complex formation in the purified system, but with reduced potency relative to doxorubicin. The type of the 3'-N-position substituent determined whether Group 3 analogues were cytotoxic and strong intercalators, or less active and nonintercalating. Although C-14-unsubstituted intercalators of Group 3 did not form cleavable complexes in the purified system, they were cytotoxic. The study shows that DNA intercalation is required but not sufficient for the activity by topo-II-targeted anthracyclines. In addition to the planar chromophore which is involved in intercalation, two other domains of the anthracycline molecule are important for the interaction with topo-II: (a) substitution of the C-14 position totally inhibits drug activity in the purified system, but enhances cytotoxicity by aiding drug uptake and presumably acting on other cellular targets; and (b) substitutions on the 3'-N position of the sugar ring can, depending on the nature of the substituent, inhibit intercalation and/or topo-II-targeting activity. These findings may provide guidance for the synthesis and development of new active analogues.     

In vitro comparative studies of the myelotoxicity and antitumor activity of 6-[bis-(2-chloroethyl)-amino]-6-deoxy-D-glucose versus melphalan utilizing the CFU-C and HTSCA assays

Summary 6-[Bis-(2-chloroethyl)-amino]-6-deoxy-D-glucose (C-6) is a new glucose-containing nitrogen mustard that has significant activity for murine P388 leukemia with relative sparing of bone marrow in mice. The in vitro myelotoxicity of C-6 compared with that of melphalan, a clinically active, myelosuppressive nitrogen mustard, was determined in the CFU-C assay in human bone marrow samples obtained from normal volunteers. There was no significant difference between the myelosuppressive actions of C-6 and melphalan at any of the concentrations used except for 4.0 M, at which C-6 was significantly (P0.05) more toxic than melphalan. Both agents decreased the number of bone marrow cell colonies to approximately 12% of control at 6.6 M (1 h incubation), which is a good approximation of melphalan's CxT (concentration by time) in man.We used the human tumor stem cell assay (HTSCA) to investigate in vitro antitumor activity. We obtained two specimens of malignant melanoma and two of malignant ovarian carcinoma from patients not previously treated with chemotherapy. The antitumor activity of melphalan was either similar to or greater than that of C-6 at all concentrations utilized against any of the four tumor specimens, except at 1.3 M for tumor I. In particular, there was no significant difference in the antitumor activities of the two agents at 6.6 M. These results suggest that C-6 will not be less myelosuppressive than melphalan at doses that produce equivalent antitumor activity in man.In addition, C-6 did not demonstrate increased myelotoxicity for normal human bone marrow cells incubated in glucose-deficient medium as against medium containing 300 mg% glucose at any of the concentrations used. This suggests that C-6 is not transported into normal human bone marrow cells via the glucose transport system, despite the presence of a glucose moiety within the molecale.Supported by Grant RO1 CA28984, a grant from the National Cancer Institute of Canada, a private contribution from Mr Schiff     

Hypoglycemia associated with carcinoid tumors. A case report and review of the literature

The authors report a 73-year-old man with a jejunal carcinoid tumor and spontaneous hypoglycemia. Plasma insulin was suppressed (insulin/glucose ratio, 0.2), and nonsuppressible insulin-like proteins (NSILP) were elevated. Seven other patients with carcinoid or related tumors associated with hypoglycemia were found in the literature. Nonsuppressible insulin-like proteins were not measured in these patients.     

A Tumour Cell Aggregation Promoting Substance from Rat Ascites Hepatoma Cells

A substance capable of promoting tumour cell aggregation was released from rat ascites hepatoma cell (possibly from the cell surface) kept in Hanks'' balanced salt solution (free of calcium and magnesium) in the cold, and then partially purified by chromatography with DEAE-Sephadex and gel filtration with Bio-gel. The thermostable substance seemed to be a glycoprotein and its molecular weight was about 72,000 when measured by gel filtration on Sephadex G-200. It had no proteolytic activity. The material was clearly effective for rat ascites hepatoma cells as well as SV40 transformed cells, but less effective for Chang''s cells and apparently ineffective for normal rat liver cells and red blood cells. The action of this material was more potent than that of Jack bean concanavalin A when assayed for aggregation of SV40 transformed cells. Its effect was not influenced by concanavalin A inhibitors such as alpha-methyl-D-glucopyranoside, N-acetyl-D-glucosamine and D-glucose.     

Antitumor activity and bone marrow toxicity of aminoglucose mustard anticancer agents in mice

In previous structure-activity studies, we have demonstrated that attachment of a glucose molecule to the chloroethylnitrosourea cytotoxic group produces a compound with reduced murine bone marrow toxicity and retention of full antitumor activity. To further define this protective role conferred by the glucose moiety in bone marrow cells, we have replaced the nitrosourea cytotoxic group with another class of alkylating agent, a bifunctional nitrogen mustard. In a detailed structure-activity analysis, we have now characterized four analogues, with the mustard cytotoxic group positioned at carbon 2 [1,3,4,6-tetra-O-acetyl-2-(di-2-chloroethyl)amino-2-deoxy-D-glucopyranos e (TGM)], carbon 6, or carbon 1 (D- and L-isomers) of the aminoglucose molecule. On a molar basis, TGM was most toxic to normal BALB/c X DBA/2 F1 mice, with a 10% lethal dose (LD10) of 3.8 mumol/kg. The D- and L-isomers of 2,3,4,6-tetra-O-acetyl-N,N-bis(2-chloroethyl)glucopyranosylamine (C-1) were the least toxic, with an LD10 of 73 mumol/kg for both. Optimal antitumor activity against the murine P388 leukemia (single i.p. administration of the LD10) did not differ significantly among the four analogues, with increased life span ranging from 83-86%. P388 antitumor activity for nitrogen mustard (HN2) was significantly less, 60% increased life span (P = 0.01), while p-di(2-chloroethyl)amino-L-phenylalanine produced an increased life span of greater than 101%. An LD10 of 6-bis-(2-chloroethyl) amino-6-deoxy-D-glucose (C-6) or TGM produced significantly less depression of WBC counts than did an equitoxic dose of the C-1 isomers, HN2, or p-di(2-chloroethyl)amino-L-phenylalanine. The mean nadir WBC count for C-6 equaled 86% of control, and for TGM, 80% of control. Consistent with this sparing effect on the peripheral WBC, C-6 and TGM produced significantly less in vivo murine bone marrow DNA synthesis depression, 77 and 64% of control, respectively, as compared to the depression nadir produced by HN2 (27% of control), the D-isomer of C-1 (17%), the L-isomer of C-1 (18%), and p-di(2-chloroethyl)amino-L-phenylalanine (2%). These structure-activity studies demonstrate that conjugation of the mustard cytotoxic group to carbon 6 or carbon 2 of glucose produces an analogue that retains P388 antitumor activity significantly greater than that of HN2, with a concomitant reduction in murine bone marrow toxicity.     

A kinase-independent biological activity for insulin growth factor-1 receptor (IGF-1R): Implications for Inhibition of the IGF-1R signal

It has been demonstrated that epidermal growth factor receptor (EGFR) can have kinase independent activity. EGFR kinase-independent function maintains intracellular glucose levels via sodium glucose transporter protein 1 (SGLT1) and supports cell survival. It is plausible that this phenomenon can apply to other receptor tyrosine kinases. We found that transfection of insulin-like growth factor receptor (IGF-1R) siRNA into HEK293 (human embryonic kidney) and MCF7 (metastatic breast cancer) cells results in decreased intracellular glucose levels, whereas treatment with the IGF-1R tyrosine kinase inhibitor OSI-906 did not affect intracellular glucose levels. In addition, IGF-1R interacted with SGLT1 in a manner similar to that previously reported with EGFR. The combination of IGF-1R siRNA and OSI-906 resulted in decreased viability of HEK293 and MCF7 cell lines compared to either agent alone. Collectively, these experiments suggest that IGF-1R, has kinase-independent biologic functions and provide a rationale for combining anti-IGF-1R antibodies or siRNA and IGF-1R small molecule inhibitors.     

Pharmacokinetics and tissue distribution of cryptophycin 52 (C-52) epoxide and cryptophycin 55 (C-55) chlorohydrin in mice with subcutaneous tumors

PURPOSE: To compare the pharmacokinetics and tissue distribution (both normal and tumor) of cryptophycin 52 (C-52) and its putative chlorohydrin prodrug cryptophycin 55 (C-55) in a murine model and to investigate a possible mechanism behind the superior activity of C-55. METHODS: Mammary adenocarcinoma 16/c tumor-bearing mice were treated with an i.v. bolus of 11 mg/kg C-52 or 38 mg/kg C-55 in Cremophor-alcohol. At predetermined time intervals, C-52 and C-55 concentrations in plasma, liver, kidney, small intestine and tumors were measured using a previously described HPLC method. Pharmacokinetic parameters were computed using noncompartmental methods. Tissue (both normal and tumor) to plasma ratios as a function of time were also calculated for comparison. RESULTS: Both C-52 and C-55 were rapidly distributed into different tissues including tumors following i.v. administration. However, the affinities of these compounds towards different tissues were different. Thus, the half-lives (minutes) of C-55 were in the decreasing order liver (725), intestine (494), tumor (206), kidney (62) and plasma (44), whereas the AUC values (microg x min/ml) were in the order tumor (9077), liver (7734), kidney (6790), plasma (2372) and intestine (2234). For C-52, the half-lives (minutes) were in the decreasing order liver (1333), kidney (718), intestine (389), tumor (181) and plasma (35), and the AUC values (microg x min/ml) were in the order kidney (1164), liver (609), intestine (487), plasma (457) and tumor (442). The relative exposures to C-52 after i.v. injection of C-55 were plasma 3.9%, tumor 80.8%, kidney 3.4%, liver 1.1% and intestine 2.8%. Although plasma exposure to C-52 following C-55 administration was relatively small, the use of C-55 to deliver C-52 increased the retention of C-52 and its AUC in tumor compared to direct injection of C-52. Simultaneously, this approach shortened C-52 retention in all normal tissues studied. CONCLUSIONS: The distribution of C-55 and its bioconversion to C-52 in different organs and tumor tissue observed in this study suggest the ability of C-55 to target tumor tissue, creating a depot of C-52 in tumor. Increased C-52 exposure of tumor, with concomitant decreased exposure of normal tissue, is a contributing factor to the superior activity of C-55 versus C-52. However, except in the case of tumor tissue in which 81% of C-55 converts to C-52, only a minor amount of C-55 may serve as a prodrug for C-52, whereas the majority is handled by the biosystem through a different route of elimination. Tissue distribution combined with rate of conversion may be an important determinant of the relative effectiveness of other epoxide-chlorohydrin pairs of cryptophycins.     

Expression of alien H-2 specificities of a chemically induced BALB/c fibrosarcoma.

The presence of alien histocompatibility antigens on the cell surface of the 3-methylcholanthrene-induced BALB/c (H-2d) fibrosarcoma C-1, was investigated by serological and transplantation studied. Absorption experiments with monospecific alloantisera showed that C-1 cells expressed their original private (H-2.4 and 31) and public (H-2.3, 8, 28, and 35) specificities. C-1 cells were also able to absorb monospecific antisera directed to the private specificity H-2.23 of the H-2k haplotype, as well as antisera to the public specificities H-2.1, 5, 11 11 and 25 (H-2k and in part H-2q, H-2a and H-2b haplotypes), which are absent from H-2d normal cells. Conversely, other alien specificities (H-2.2, 17, 30, 32, and 33) were not detected on C-1 cells. The C-1 cells were also unable to absorb the activity of an anti-Ia serum (1-28) directed to 1a.1, 2 and 19 (lak) specificities. Transplantation studies showed that resistance against the challenge of C-1 cells could be induced in syngeneic BALB/c mice by preimmunization with normal tissues from C3Hf and AKR (H-2k), A (H-2a) and C57BL/6J (H-2b) strains (expressing all or some of the extra H-2 antigens of the tumor) whereas no protection was obtained with DBA/2 (H-2d) or with W/Fu rat tissues. The anti-tumor activity could be passively transferred by BALB/c lymphoid cells immune to normal C3Hf, AKR, A, and NIH (H-2q) tissues, but no protection was achieved with lymphoid cells immune to DBA/2 or to W/Fu normal rat tissues. These data indicate that foreign H-2 antigens are expressed on C-1 tumor and that they might function as tumor-associated transplantation antigen which was shown to be present and individually distinct on this sarcoma by appropriate in vivo tests.     

Increased Activity of Insulin-like Growth Factor-binding Protein in Human Thyroid Papillary Cancer Tissue

It has been shown that both insulin-like growth factor-I (IGF-I) and IGF-binding proteins (IGFBPs) are produced by thyroid cells in culture and that the cells respond to IGF-I with increased DNA synthesis, suggesting an autocrine/paracrine role of IGF-I in the regulation of thyroid cell growth. We investigated the tissue contents of immunoreactive IGF-I (irIGF-I) and IGFBPs in human papillary carcinoma and compared them with those of normal thyroid tissue. When irIGF-I was measured after separation of the IGFBPs by gel-filtration, its content in carcinoma tissue was not different from that in adjacent normal tissue (566±58 vs. 424±75 pg/mg protein, N = 10). Nor was there any difference in the abundance of IGF-I mRNA expression determined by slot blot analysis. On the other hand, IGFBP activity measured in terms of ¹²⁵I-IGF-I binding was significantly higher in cancer extracts. Western ligand blot analysis of IGFBPs revealed several species (24–42 kDa) of IGFBPs. The IGF-I-binding activity of 38–41 kDa species (corresponding to IGFBP-3) was not different between extracts of cancer tissue and those of normal tissue, whereas that of 28–32 kDa species was significantly higher in cancer tissue extracts. Since IGFBPs have been reported to modulate cellular responses to IGF-I, the present data suggest that higher IGFBP activity in cancer tissue is involved in regulating growth of thyroid papillary carcinoma cells.     

Metabolic effects of tumour necrosis factor alpha in NMRI mice

Following a single injection of 7.5 x 10(7) U kg-1 of human recombinant tumour necrosis factor-alpha (TNF-alpha) to female NMRI mice, marked hypoglycaemia was observed within a 2 h period, accompanied by a severe depletion of liver glycogen and a drop in rectal body temperature when compared with pair-fed controls. There was no alteration in plasma alanine, lactate or pyruvate values, but an elevation of acetoacetate and 3-hydroxybutyrate when compared with pair-fed controls. Production of 14CO2 from U-14C-glucose was reduced in TNF-alpha treated animals, while production of 14CO2 from U-14C-palmitate was not significantly different from controls, suggesting that the glucose was not being used to provide an increased metabolic rate. Glucose utilisation by different tissues was investigated by the 2-deoxyglucose tracer method. This showed that 2 h following TNF-alpha infusion glucose utilisation was increased in colon, liver, kidney and spleen by 500, 350, 36 and 25% respectively. However, when calculated on a whole-animal basis the major contributor to the increased glucose consumption was the liver. Plasma levels of both FFA and triglycerides were also elevated in TNF-alpha treated animals, suggesting that increased consumption of glucose by the liver may be utilised for lipogenesis. The rate of conversion of glucose into lipids in the liver was more than doubled 2 h after TNF-alpha administration with a concomitant rise in plasma and adipose tissue. These results suggest that administration of TNF-alpha produces a severe hypoglycaemia in order to serve an increased lipogenesis in liver and adipose tissue, which appears to be independent of the anorectic effect.     

Functional and biochemical parameters of activation related to macrophage cytostatic effects on tumor cells

The cytostatic action of non-immune, activated peritoneal rat macrophages (AM) on tumor targets largely depends upon the duration of their culture in vitro. AM show full cytostatic activity for the first 12-24 h of culture followed by gradual loss. This distinctive pattern of cytostatic activity displayed by AM was compared with the course of a variety of functional, metabolic and biochemical parameters. DNase, RNase, esterase, acid phosphatase, glucose oxidation and cellular adherence remained unchanged or decreased whereas prolonged culture led to an increase in activity of other lysosomal enzymes. Pinocytosis and phagocytosis were most marked in macrophages between 12-60 h whereas elevation in macrophage protein synthesis was high only after 4 to 12 h of culture, followed by a gradual decline. Thus the capacity of macrophages to stop proliferation of tumor targets closely parallels the level of protein synthesis, whereas other parameters of macrophage function such as lysosomal enzymes, glucose oxidation, cellular adherence, pinocytosis and phagocytosis, manifested a clearly different timecourse.     

L-carnitine ameliorates cancer cachexia in mice by regulating the expression and activity of carnitine palmityl transferase

Cancer cachexia is characterized by progressive weight loss with the depletion of adipose tissue and skeletal muscle. Impaired fatty acid oxidation mainly resulting from the decrease of carnitine palmitoyltransferase I and II activities in the liver is an important factor that contributes to cancer cachexia . Although recent studies suggest a potential application of L-carnitine in treatment of cancer cachexia, the underlying mechanisms are unknown. In the present study, we aim to assess the effects of L-carnitine on the activity and expression of CPT I and II in the liver of cachectic cancer mice. Our results show that the inoculation of colon-26 adenocarcinoma cells into mice led to cancer cachexia characterized by notable decreases in food intake, gastrocnemius muscle and epididymus fat weight. In addition, the mRNA level and activity of liver carnitine palmitoyltransferase (CPT) I and II, and serum levels of free carnitine and acetylcarnitine were markedly decreased in cachectic mice, accompanied by marked increases in serum levels of tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6). A continuous oral treatment with L-carnitine at 18 mg/kg per day increased dietary uptake, gastrocnemius muscle weight and epididymus fat weight, increased blood glucose and serum albumin levels, and decreased total cholesterol level in cancer cachectic mice, but did not affect tumor growth. These effects of L-carnitine on cancer cachexia mice were accompanied by the upregulation of mRNA level of CPT I and II and increased enzyme activity of CPT I in the liver, as well as the downregulation of serum TNF-α and IL-6 levels. Moreover, free carnitine levels were negatively correlated with serum TNF-α or IL-6 level. These results indicate that L-carnitine ameliorates cancer cachexia by regulating serum TNF-α and IL-6 levels and modulating the expression and activity of CPT in the liver.