Aurelia Lugea

The pancreatic stellate cell: a star on the rise in pancreatic diseases

Pancreatic stellate cells (PaSCs) are myofibroblast-like cells found in the areas of the pancreas that have exocrine function. PaSCs are regulated by autocrine and paracrine stimuli and share many features with their hepatic counterparts, studies of which have helped further our understanding of PaSC biology. Activation of PaSCs induces them to proliferate, to migrate to sites of tissue damage, to contract and possibly phagocytose, and to synthesize ECM components to promote tissue repair. Sustained activation of PaSCs has an increasingly appreciated role in the fibrosis that is associated with chronic pancreatitis and with pancreatic cancer. Therefore, understanding the biology of PaSCs offers potential therapeutic targets for the treatment and prevention of these diseases.

Cell Death in Pancreatitis CASPASES PROTECT FROM NECROTIZING PANCREATITIS

Mechanisms of cell death in pancreatitis remain unknown. Parenchymal necrosis is a major complication of pancreatitis; also, the severity of experimental pancreatitis correlates directly with necrosis and inversely with apoptosis. Thus, shifting death responses from necrosis to apoptosis may have a therapeutic value. To determine cell death pathways in pancreatitis and the possibility of necrosis/apoptosis switch, we utilized the differences between the rat model of cerulein pancreatitis, with relatively high apoptosis and low necrosis, and the mouse model, with little apoptosis and high necrosis. We found that caspases were greatly activated during cerulein pancreatitis in the rat but not mouse. Endogenous caspase inhibitor X-linked inhibitor of apoptosis protein (XIAP) underwent complete degradation in the rat but remained intact in the mouse model. Furthermore, XIAP inhibition with embelin triggered caspase activation in the mouse model, implicating XIAP in caspase blockade in pancreatitis. Caspase inhibitors decreased apoptosis and markedly stimulated necrosis in the rat model, worsening pancreatitis parameters. Conversely, caspase induction with embelin stimulated apoptosis and decreased necrosis in mouse model. Thus, caspases not only mediate apoptosis but also protect from necrosis in pancreatitis. One protective mechanism is through degradation of receptor-interacting protein (RIP), a key mediator of “programmed” necrosis. We found that RIP was cleaved (i.e. inactivated) in the rat but not the mouse model. Caspase inhibition restored RIP levels; conversely, caspase induction with embelin triggered RIP cleavage. Our results indicate key roles for caspases, XIAP, and RIP in the regulation of cell death in pancreatitis. Manipulating these signals to change the pattern of death responses presents a therapeutic strategy for treatment of pancreatitis.

A Starring Role for Stellate Cells in the Pancreatic Cancer Microenvironment

Pancreatic ductal adenocarcinoma is a devastating disease, and patient outcomes have not improved in decades. Treatments that target tumor cells have largely failed. This could be because research has focused on cancer cells and the influence of the stroma on tumor progression has been largely ignored. The focus of pancreatic cancer research began to change with the identification of pancreatic stellate cells, which produce the pancreatic tumor stroma. There is compelling in vitro and in vivo evidence for the influence of pancreatic stellate cells on pancreatic cancer development; several recent preclinical studies have reported encouraging results with approaches designed to target pancreatic stellate cells and the stroma. We review the background and recent advances in these areas, along with important areas of future research that could improve therapy.

Desmoplasia of Pancreatic Ductal Adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) is the most common form of pancreatic cancer and is characterized by remarkable desmoplasia. The desmoplasia is composed of extracellular matrix (ECM) proteins, myofibroblastic pancreatic stellate cells, and immune cells associated with a multitude of cytokines, growth factors, and ECM metabolizing enzymes. The mechanisms of participation of this complex matrix process in carcinogenesis are only starting to be appreciated. Recent studies showed key roles for stellate cells in the production of ECM proteins as well as cytokines and growth factors that promote the growth of the cancer cells all present in the desmoplastic parts of PDAC. In addition, interactions of ECM proteins and desmoplastic secreted growth factors with the cancer cells of PDAC activate intracellular signals including reactive oxygen species that act to make the cancer cells resistant to dying. These findings suggest that the desmoplasia of PDAC is a key factor in regulating carcinogenesis of PDAC as well as responses to therapies. A better understanding of the biology of desmoplasia in the mechanism of PDAC will likely provide significant opportunities for better treatments for this devastating cancer.

Synergistic steatohepatitis by moderate obesity and alcohol in mice despite increased adiponectin and p-AMPK

BACKGROUND & AIMS Mechanisms underlying synergistic liver injury caused by alcohol and obesity are not clear. We have produced a mouse model of synergistic steatohepatitis by recapitulating the natural history of the synergism seen in patients for mechanistic studies. METHODS Moderate obesity was induced in mice by 170% overnutrition in calories using intragastric overfeeding of high fat diet. Alcohol (low or high dose) was then co-administrated to determine its effects. RESULTS Moderate obesity plus alcohol intake causes synergistic steatohepatitis in an alcohol dose-dependent manner. A heightened synergism is observed when a high alcohol dose (32g/kg/d) is used, resulting in plasma ALT reaching 392±28U/L, severe steatohepatitis with pericellular fibrosis, marked M1 macrophage activation, a 40-fold induction of iNos, and intensified nitrosative stress in the liver. Hepatic expression of genes for mitochondrial biogenesis and metabolism are significantly downregulated, and hepatic ATP level is decreased. Synergistic ER stress evident by elevated XBP-1, GRP78 and CHOP is accompanied by hyperhomocysteinemia. Despite increased caspase 3/7 cleavage, their activities are decreased in a redox-dependent manner. Neither increased PARP cleavage nor TUNEL positive hepatocytes are found, suggesting a shift of apoptosis to necrosis. Surprisingly, the synergism mice have increased plasma adiponectin and hepatic p-AMPK, but adiponectin resistance is shown downstream of p-AMPK. CONCLUSIONS Nitrosative stress mediated by M1 macrophage activation, adiponectin resistance, and accentuated ER and mitochondrial stress underlie potential mechanisms for synergistic steatohepatitis caused by moderate obesity and alcohol.

Adaptive unfolded protein response attenuates alcohol-induced pancreatic damage.

BACKGROUND & AIMS Endoplasmic reticulum (ER) stress responses (collectively known the unfolded protein response [UPR]) have important roles in several human disorders, but their contribution to alcoholic pancreatitis is not known. We investigated the role of X-box binding protein 1 (XBP1), a UPR regulator, in prevention of alcohol-induced ER stress in the exocrine pancreas. METHODS Wild-type and Xbp1(+/-) mice were fed control or ethanol diets for 4 weeks. Pancreatic tissue samples were then examined by light and electron microscopy to determine pancreatic alterations; UPR regulators were analyzed biochemically. RESULTS In wild-type mice, ethanol activated a UPR, increasing pancreatic levels of XBP1 and XBP1 targets such as protein disulfide isomerase (PDI). In these mice, pancreatic damage was minor. In ethanol-fed Xbp1(+/-) mice, XBP1 and PDI levels were significantly lower than in ethanol-fed wild-type mice. The combination of XBP1 deficiency and ethanol feeding reduced expression of regulators of ER function and the up-regulation of proapoptotic signals. Moreover, ethanol feeding induced oxidation of PDI, which might compromise PDI-mediated disulfide bond formation during ER protein folding. In ethanol-fed Xbp1(+/-) mice, ER stress was associated with disorganized and dilated ER, loss of zymogen granules, accumulation of autophagic vacuoles, and increased acinar cell death. CONCLUSIONS Long-term ethanol feeding causes oxidative ER stress, which activates a UPR and increases XBP1 levels and activity. A defective UPR due to XBP1 deficiency results in ER dysfunction and acinar cell pathology.

Epidemiology, risk factors, and the promotion of pancreatic cancer: Role of the stellate cell

There are approximately 277 000 new cases of pancreatic cancer and 266 000 deaths from pancreatic cancer annually, indicating a mortality rate of 96% of the cases diagnosed. Because of the ineffectiveness of therapies, a major emphasis needs to be placed on prevention. This paper reviews the epidemiology and risk factors for pancreatic cancer, and uses this information to propose plausible research directions for determining the biological mechanisms mediating the effects of risk factors on the promotion of pancreatic cancer, with a focus on the pancreatic stellate cell.

High-Fat, High-Calorie Diet Promotes Early Pancreatic Neoplasia in the Conditional KrasG12D Mouse Model

There is epidemiologic evidence that obesity increases the risk of cancers. Several underlying mechanisms, including inflammation and insulin resistance, are proposed. However, the driving mechanisms in pancreatic cancer are poorly understood. The goal of the present study was to develop a model of diet-induced obesity and pancreatic cancer development in a state-of-the-art mouse model, which resembles important clinical features of human obesity, for example, weight gain and metabolic disturbances. Offspring of Pdx-1-Cre and LSL-KrasG12D mice were allocated to either a high-fat, high-calorie diet (HFCD; ∼4,535 kcal/kg; 40% of calories from fats) or control diet (∼3,725 kcal/kg; 12% of calories from fats) for 3 months. Compared with control animals, mice fed with the HFCD significantly gained more weight and developed hyperinsulinemia, hyperglycemia, hyperleptinemia, and elevated levels of insulin-like growth factor I (IGF-I). The pancreas of HFCD-fed animals showed robust signs of inflammation with increased numbers of infiltrating inflammatory cells (macrophages and T cells), elevated levels of several cytokines and chemokines, increased stromal fibrosis, and more advanced PanIN lesions. Our results show that a diet high in fats and calories leads to obesity and metabolic disturbances similar to humans and accelerates early pancreatic neoplasia in the conditional KrasG12D mouse model. This model and findings will provide the basis for more robust studies attempting to unravel the mechanisms underlying the cancer-promoting properties of obesity, as well as to evaluate dietary- and chemopreventive strategies targeting obesity-associated pancreatic cancer development. Cancer Prev Res; 6(10); 1064–73. ©2013 AACR.

Alcohol Abuse, Endoplasmic Reticulum Stress and Pancreatitis

Alcohol abuse is a common cause of both acute and chronic pancreatitis. There is a wide spectrum of pancreatic manifestations in heavy drinkers from no apparent disease in most individuals to acute inflammatory and necrotizing pancreatitis in a minority of individuals with some progressing to chronic pancreatitis characterized by replacement of the gland by fibrosis and chronic inflammation. Both smoking and African-American ethnicity are associated with increased risk of alcoholic pancreatitis. In this review we describe how our recent studies demonstrate that ethanol feeding in rodents causes oxidative stress in the endoplasmic reticulum (ER) of the digestive enzyme synthesizing acinar cell of the exocrine pancreas. This ER stress is attenuated by a robust unfolded protein response (UPR) involving X-box binding protein-1 (XBP1) in the acinar cell. When the UPR activation is prevented by genetic reduction in XBP1, ethanol feeding causes significant pathological responses in the pancreas. These results suggest that the reason most individuals who drink alcohol heavily do not get significant pancreatic disease is because the pancreas mounts an adaptive UPR to attenuate the ER stress that ethanol causes. We hypothesize that disease in the pancreas results when the UPR is insufficiently robust to alleviate the ER stress caused by alcohol abuse.

Animal and in vitro models of alcoholic pancreatitis: role of cholecystokinin.

Although ethanol abuse is the major etiologic factor in the development of acute and chronic pancreatitis, the mechanisms of ethanol effects to cause pancreatitis are poorly understood. The major reason for the lack of progress is the relative lack of animal models that reproduce the deleterious effects of ethanol on the pancreas that are observed in human disease. We propose that the effect of ethanol on the pancreas is due to its ability to sensitize animals and humans to the potentially injurious effects of other stimuli. We have developed models of ethanol-induced acute and chronic pancreatitis in rats as well as pancreatic acinar cells in primary culture demonstrating that ethanol sensitizes the pancreas to the inflammatory, cell death, and fibrosing responses caused by cholecystokinin (CCK). Our results indicate that the ethanol-sensitized inflammatory response is the key or trigger event for the development of the other pathologic responses in both acute and chronic pancreatitis, such as cell death, intracellular digestive enzyme activation, and fibrosis. These findings suggest that experimental strategies designed to reveal the modulating effects of ethanol on the mechanisms underlying the inflammatory, cell death, and fibrosing responses stimulated by CCK will provide the key information needed to understand how ethanol abuse causes pancreatitis.