Interleukin-4 (IL-4) is an immunomodulatory cytokine, which can inhibit the growth of tumour cells. Pancreatic cancer cells and tissues express high levels of IL-4 receptors. The aim of this study was to characterise the effects of IL-4 on the growth and signalling pathways of pancreatic cancer cells. Cell growth was determined by cell counting and MTT assays in association with fluorescence-activated cell sorter analysis, IL-4 expression using ELISA and real-time PCR techniques, and signal transduction using immunoprecipitation or immunoblot analysis. We now report for the first time that IL-4 significantly enhanced the growth of five out of six cultured pancreatic cancer cell lines in a dose-dependent manner in association with an increased fraction of cells in S-phase. Surprisingly, all six cell lines expressed endogenous IL-4, and IL-4 was detectable in the supernatant. Incubating cells with neutralising IL-4 antibodies resulted in a significant inhibition of basal growth in three cell lines, including IL-4-unresponsive MIA PaCa-2 cells, which however expressed the highest endogenous IL-4 levels. Interleukin-4 enhanced activity of MAPK, Akt-1, and Stat3 in IL-4-responsive, but not in IL-4-unresponsive MIA PaCa-2 cells; however, IL-4 enhanced tyrosine phosphorylation of insulin receptor substrate-1 and -2 in all cell lines. Our results demonstrate for the first time that pancreatic cancer cells produce IL-4 and that IL-4 can act as a growth factor in pancreatic cancer cells. Together with the observation that neutralising IL-4 antibodies can inhibit the growth of these cells, our results suggest that IL-4 may act as an autocrine growth factor in pancreatic cancer cells and also give rise to the possibility that cancer-derived IL-4 may suppress cancer-directed immunosurveillance in vivo in addition to its growth-promoting effects, thereby facilitating pancreatic tumour growth and metastasis.

Pancreatic ductal adenocarcinoma (PDAC) metastasizes to liver at early stages, making this disease highly lethal. Tissue inhibitor of metalloproteinases-1 (TIMP1) creates a metastasis-susceptible environment in the liver. We investigated the role of TIMP1 and its receptor CD63 in metastasis of early-stage pancreatic tumors using mice and human cell lines and tissue samples. We obtained liver and plasma samples from patients in Germany with chronic pancreatitis, pancreatic intra-epithelial neoplasia, or PDAC, as well as hepatic stellate cells (HSCs). We performed studies with Ptf1a⁺/Cre;Kras⁺/LSL-G12D;Trp53loxP/loxP (CPK) mice, Pdx-1⁺/Cre;Kras⁺/LSL-G12D;Trp53⁺/LSL-R172H (KPC) mice, and their respective healthy littermates as control, and Cd63^−/− mice with their wild-type littermates. KPC mice were bred with Timp1^−/− mice to produce KPCxTimp1^−/− mice. TIMP1 was overexpressed and CD63 was knocked down in mice using adenoviral vectors AdTIMP1 or AdshCD63, respectively. Hepatic susceptibility to metastases was determined after intravenous inoculation of syngeneic 9801L pancreas carcinoma cells. Pancreata and liver tissues were collected and analyzed by histology, immunohistochemical, immunoblot, enzyme-linked immunosorbent assay, and quantitative polymerase chain reaction analyses. We analyzed the effects of TIMP1 overexpression or knockdown and CD63 knockdown in transduced human primary HSCs and HSC cell lines. Chronic pancreatitis, pancreatic intra-epithelial neoplasia, and PDAC tissues from patients expressed higher levels of TIMP1 protein than normal pancreas. The premalignant pancreatic lesions that developed in KPC and CPK mice expressed TIMP1 and secreted it into the circulation. In vitro and in vivo, TIMP1 activated human or mouse HSCs, which required interaction between TIMP1 and CD63 and signaling via phosphatidylinositol 3-kinase, but not TIMP1 protease inhibitor activity. This signaling pathway induced expression of endogenous TIMP1. TIMP1 knockdown in HSCs reduced their activation. Cultured TIMP1-activated human and mouse HSCs began to express stromal-derived factor-1, which induced neutrophil migration, a marker of the premetastatic niche. Mice with pancreatic intra-epithelial neoplasia–derived systemic increases in TIMP1 developed more liver metastases after injections of pancreatic cancer cells than mice without increased levels of TIMP1. This increase in formation of liver metastases from injected pancreatic cancer cells was not observed in TIMP1 or CD63 knockout mice. Expression of TIMP1 is increased in chronic pancreatitis, pancreatic intra-epithelial neoplasia, and PDAC tissues from patients. TIMP1 signaling via CD63 leads to activation of HSCs, which create an environment in the liver that increases its susceptibility to pancreatic tumor cells. Strategies to block TIMP1 signaling via CD63 might be developed to prevent PDAC metastasis to the liver.

Cachexia is a multi-factorial, systemic syndrome that especially affects patients with cancer of the gastrointestinal tract, and leads to reduced treatment response, survival and quality of life. The most important clinical feature of cachexia is the excessive wasting of skeletal muscle mass. Currently, an effective treatment is still lacking and the search for therapeutic targets continues. Even though a substantial number of animal studies have contributed to a better understanding of the underlying mechanisms of the loss of skeletal muscle mass, subsequent clinical trials of potential new drugs have not yet yielded any effective treatment for cancer cachexia. Therefore, we questioned to which degree findings from animal studies can be translated to humans in clinical practice and research. A substantial amount of animal studies on the molecular mechanisms of muscle wasting in cancer cachexia has been conducted in recent years. This extensive review of the literature showed that most of their observations could not be consistently reproduced in studies on human skeletal muscle samples. However, studies on human material are scarce and limited in patient numbers and homogeneity. Therefore, their results have to be interpreted critically. More research is needed on human tissue samples to clarify the signaling pathways that lead to skeletal muscle loss, and to confirm pre-selected drug targets from animal models in clinical trials. In addition, improved diagnostic tools and standardized clinical criteria for cancer cachexia are needed to conduct standardized, randomized controlled trials of potential drug candidates in the future.

To address possible effects of heat shock protein 70 (Hsp70) on energy metabolism, we established a cell line expressing different levels of Hsp70 and evaluated changes in glucose and lactate metabolites, as well as ATP levels accordingly. In addition, activities of enzymes involved in glycolysis [phosphofructokinase (PFK) and lactate dehydrogenase (LDH)], Krebs cycle [citric synthase (CS)], and oxidative phosphorylation {NADH dehydrogenase [complex I (CI)] and ubiquinol:cytochrome-c reductase [complex III (CIII)]} were analyzed. The results show that both glucose consumption and lactate excretion were elevated significantly in cells expressing increased levels of Hsp70. Simultaneously, the activities of glycolytic enzymes PFK and LDH were increased markedly in cells overexpressing Hsp70. Activities of enzymes CI and CIII, both involved in oxidative phosphorylation, decreased upon increased expression of Hsp70. These findings were supported by nonsignificant reductions of CS activities in cells that overexpressed Hsp70, whereas intracellular ATP levels remained constant over a wide range of Hsp70 expression. In conclusion, overexpression of Hsp70 in HeLa cells results in downregulation of oxidative phosphorylation, in particular, multiprotein CIII, the main source of reactive oxygen species. In exchange, upregulation of the glycolytic pathway compensates for the homeostasis of cellular ATP supply.

Sex disparity in cancer is so far inadequately considered, and components of its basis are rather unknown. We reveal that male versus female pancreatic cancer (PC) patients and mice show shortened survival, more frequent liver metastasis, and elevated hepatic metastasis-promoting gene expression. Tissue inhibitor of metalloproteinases 1 (TIMP1) was the secreted factor with the strongest male-biased expression in patient-derived pancreatic tumors. Male-specific up-regulation of systemic TIMP1 was demonstrated in PC mouse models and patients. Using TIMP1-competent and TIMP1-deficient PC mouse models, we established a causal role of TIMP1 in determining shortened survival and increased liver metastasis in males. Observing TIMP1 expression as a risk parameter in males led to identification of a subpopulation exhibiting increased TIMP1 levels (T1HI males) in both primary tumors and blood. T1HI males showed increased risk for liver metastasis development not only in PC but also in colorectal cancer and melanoma. This study reveals a lifestyle-independent sex disparity in liver metastasis and may open new avenues toward precision medicine.

During the acute training response, peripheral cellular mechanisms are mainly metabolostatic to achieve energy supply. During prolonged training, glycogen deficiency occurs; this is associated with increased expression of local cytokines, and decreased insulin secretion and β-adrenergic stimulation and lipolysis in adipose tissue which looses energy. This is indicated by decrease of adipocyte hormone leptin, which has inhibitory effects on excitatory hypothalamic neurons. Leptin, insulin, and cytokines such as interleukin 6 (IL-6) contribute to the metabolic error signal to the hypothalamus which result in decrease of hypothalamic release hormones and sympathoadrenergic stimulation. Thyroid stimulating hormone (TSH) is correlated to the metabolic hormones leptin and insulin, and may be used as indicator of metabolic control. Because the hypothalamus integrates various error signals (metabolic, hormonal, sensory afferents, and central stimuli), the pituitary’s releasing hormones represent the functional status of an athlete. Long-term overtraining will lead to downregulation of hypothalamic hormonal and sympathoadrenergic responses, catabolism, and fatigue. These changes contribute to myopathy with predominant expression of slow muscle fiber type and inadequacy in performance. Thyroid hormones are closely involved in the training response and metabolic control.