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Pancreatic cancer remains one of the most difficult to treat human cancers despite recent advances in targeted therapy. Inhibition of isoprenylcysteine carboxylmethyltransferase (ICMT), an enzyme that posttranslationally modifies a group of proteins including several small GTPases, suppresses proliferation of some human cancer cells. However, the efficacy of ICMT inhibition on human pancreatic cancer has not been evaluated. In this study, we have evaluated a panel of human pancreatic cancer cell lines and identified those that are sensitive to ICMT inhibition. In these cells, ICMT suppression inhibited proliferation and induced apoptosis. This responsiveness to ICMT inhibition was confirmed in in vivo xenograft tumor mouse models using both a small-molecule inhibitor and shRNA-targeting ICMT. Mechanistically, we found that, in sensitive pancreatic cancer cells, ICMT inhibition induced mitochondrial respiratory deficiency and cellular energy depletion, leading to significant upregulation of p21. Furthermore, we characterized the role of p21 as a regulator and coordinator of cell signaling that responds to cell energy depletion. Apoptosis, but not autophagy, that is induced via p21-activated BNIP3 expression accounts for the efficacy of ICMT inhibition in sensitive pancreatic cancer cells in both in vitro and in vivo models. In contrast, cells resistant to ICMT inhibition demonstrated no mitochondria dysfunction or p21 signaling changes under ICMT suppression. These findings not only identify pancreatic cancers as potential therapeutic targets for ICMT suppression but also provide an avenue for identifying those subtypes that would be most responsive to agents targeting this critical enzyme. Mol Cancer Ther; 16(5); 914–23. ©2017 AACR.

Supplementary Fig. S1. ICMT inhibitor treatment leads to reduced viability in sensitive pancreatic cancer cell lines. (A) MTS viability assay that measures the total viable cells (OD measured at 490 nm) at the time of initiation (0 hr) and end (48 hr) of cysmethynil treatment for multiple pancreatic cell lines. (B) IC50 of different cell lines in response to cysmethynil treatment, determined by MTS assay shown in (A). Supplementary Fig. S2. ICMT inhibitor induces p21 and LC3 expression independent of p53. (A) Immunoblot analysis of the indicated proteins in Miapaca2 cells after treatment with 0, 20 or 22.5 μM cysmethynil for 48 h. (B) Immunoblot analysis of the indicated proteins in Miapaca2 cells with (si�p53) or without (si�control) p53 knockdown, and concurrent treatment with 0, 20, 22 and 24 μM Cysmethynil. Supplementary Fig. S3. Forced expression of BCL�XL rescues MiaPaca2 cells from apoptotic cell death induced by ICMT inhibition. (A) Light microscopy analysis of MiaPaca2 cells infected with either control retrovirus or that expressing BCL�XL, with (+) or without (�) treatment with 22.5 μM Cysmethynil for 48 h. (B) Flow cytometry assessment of the cell preparations from (A). Supplementary Fig. S4. Immunoblot study on the cell lysates prepared from AsPC1 and BxPC3 cells after 48 h treatment by either DMSO control or 22.5 μM of cysmethynil.

Metformin has been extensively studied for its impact on cancer cell metabolism and anticancer potential. Despite evidence of significant reduction in cancer occurrence in diabetic patients taking metformin, phase II cancer trials of the agent have been disappointing, quite possibly because of the lack of molecular mechanism-based patient stratification. In an effort to identify cancers that are responsive to metformin, we discovered that mitochondria respiratory capacity and respiratory reserve, which vary widely among cancer cells, correlate strongly to metformin sensitivity in both the in vitro and in vivo settings. A causal relationship between respiratory function and metformin sensitivity is demonstrated in studies in which we lowered respiratory capacity by either genetic knockdown or pharmacologic suppression of electron transport chain components, rendering cancer cells more vulnerable to metformin. These findings led us to predict, and experimentally validate, that metformin and AMP kinase inhibition synergistically suppress cancer cell proliferation.

Body mass regulation may be appeared regional differences, in order to investigate the physiological and behavioral changes in Eothenomys miletus from Shanggelila (XGLL) and Jianchuan (JC) under food restriction (FR) and refeeding (Re), body mass, food intake, resting metabolic rate (RMR), serum leptin levels, hypothalamic neuropeptides expression and activity behavior were measured. The results showed that areas and FR had significant effects on body mass, food intake, RMR, activity behavior, serum leptin levels, hypothalamic neuropeptides expression, masses of liver and small intestine in E. miletus. Body mass and serum leptin levels in XGLL were lower than that of JC, food intake, activity behavior, liver mass and RMR in XGLL were higher relative to in JC. All the indexes of the two areas of E. miletus can be restored to control levels after refeeding, showing phenotypic plasticity. In conclusion, physiological and behavioral characteristics illustrated that the influences of food and different regions on phenotypic plasticity, which had important significance for in-depth understanding of the survival and adaptation strategies of E. miletus in Hengduan mountain regions.

In the present study, liquid chromatography-mass spectrometer (LC-MS) was used to perform untargeted metabolomics analysis of white adipose tissue (WAT) and brown adipose tissue (BAT) in Tupaia belangeri during cold acclimation. Differences in biochemical composition between WAT and BAT were compared. Clarifying how the two adipose tissues respond to the lower temperature in terms of metabolomics, which elucidate the metabolic process and energy homeostasis regulation mechanism in T. belangeri. The results showed that there were 34, 59 and 20 differential metabolites in the WAT, BAT and WAT compared with BAT, respectively. WAT and BAT had significant differences in various metabolic pathways such as sugar metabolism, amino acid metabolism, lipid metabolism, and nucleotide metabolism, which were closely related to the different biological roles of the two tissues. Increasing the concentrations of intermediate products of tricarboxylic acid (TCA) cycle, pyruvic acid, and phosphoenolpyruvic acid (PEP) in WAT and increasing the metabolites in TCA cycle, glyoxylate and dicarboxylate metabolism pathways in BAT, likely to increase the thermogenic capacity in T. belangeri in response to cold stress. There were more differential metabolic pathways in BAT during cold acclimation than that of in WAT. Moreover, compared to WAT, BAT responds to cold stress by adjusting the concentration of nucleotide metabolites.

To investigate adaptive strategies of Tupaia belangeri to environmental factors in different populations, 12 locations were selected, including higher and lower altitude areas. Total of 96 and 90 metabolites were annotated in serum and liver respectively, which were mainly concentrated in primary metabolites. Principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA) showed that the locations were divided into two groups in serum metabolites, but each group had a few samples overlap. The samples of each group overlap to some degree in the liver metabolites. The tricarboxylic acid (TCA) cycle occupies a central position in metabolism. The concentrations of TCA intermediates, lipid metabolites and amino acid metabolites were higher in higher altitude areas, and the concentrations of carbohydrate and glycolysis intermediates were higher in lower altitude areas. Different areas adapted to the changes of environmental and altitude by regulating the concentration of metabolites in serum and liver, and revealed the adaptive mechanism of T. belangeri in different living environments.

This study investigated the browning plasticity of white adipose tissue (WAT) in Tupaia belangeri during cold acclimation and rewarming in order to demonstrate the adaptation mechanism of tree shrews to environmental change. The experimental group was transferred to a cold temperature, 5 ± 1 °C, acclimated for 28 d, and then returned to 25 ± 1 °C for 28 d, while the control group was maintained at the acclimation temperature, 25 ± 1 °C, for 56 d. Body mass, food intake, resting metabolic rate (RMR), WAT mass, morphology and related gene expression in male T. belangeri were measured. The results showed that body mass, food intake and RMR increased significantly under cold acclimation. There was also a significant increase in WAT mass and expression of peroxisome proliferation receptor α (PPARα), peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α), cyclooxygenaseII (COXII), bone morphogenetic protein 7 (BMP7) and the PR domain containing 16 (PRDM16), all of which decreased to control levels after rewarming. Further, WAT cells showed more multilocule adipocytes during cold acclimation, which returned to control levels after rewarming. These results suggest that browning may appear in the WAT of T. belangeri during cold acclimation. The return to control levels of WAT cell characteristics and expression of the genes involved in WAT browning after rewarming demonstrates strong browning plasticity.

Longevity, an important life-history trait, is determined by extrinsic and/or intrinsic causing mortality. Here, we used body mass (BM), field metabolic rate (FMR), longevity, and female maturity data reported from 300 amniote species to test whether 1) longevity was related to BM, FMR and female maturity, and 2) FMR, female maturity, or both, had a direct effect on longevity and whether an indirect effect of FMR on female maturity improved model fit. The results showed that BM was positively correlated with longevity and FMR, but negatively correlated with mass-specific FMR (mFMR) in amniotes. Phylogenetic confirmatory path analysis showed that, in the best model, longevity had a direct negative correlation with mFMR in lizards, and an indirect negative correlation with mFMR through female maturity in mammals. However, longevity had a direct positive correlation with mFMR in birds. Furthermore, longevity was positively correlated with female maturity in endotherms (birds and mammals) but weakly correlated with female maturity in ectotherms (lizards). Thus, our results are consistent with the life-history theory and the “rate-of-living” hypothesis in lizards and mammals but not support them in birds.

Phenotypic plasticity is the capacity for individuals to modify their phenotype in response to environmental changes. For example, variation in external and cranial characters have been documented in some small mammals and can result in controversy over classification, which has been the case for tree shrews for decades. However, there are few reports about external and cranial character variation in tree shrews (Tupaia belangeri). Here, such character variants of tree shrews were documented for 11 different sampling sites (Hainan, Daxin, Leye, Xingyi, Hekou, Kunming, Xichang, Dali, Pianma, Tengchong, Mengla) and for laboratory samples. The results showed that external and cranial characters of tree shrews significantly differed among regions, Hainan and Daxin populations were significantly larger than those of other populations, and Leye, Xingyi, Kunming and Xichang populations were smaller. The variation may be related to the habitat of tree shrews and our data may initiate new interest in morphological variants in small mammals. The presented detailed examination of the morphometric characters in tree shrews, and the exposed significant morphological differences among tree shrews, are expected to help in confirming their classification status.

The aim of this study was to examine relationships between sustainable energy intake (SusEI) and resting metabolic rate (RMR) in Eothenomys miletus and to understand the factors limiting maximum SusEI/RMR during lactation. Body mass, food intake, RMR, milk energy output (MEO), litter size, litter mass, and thermal conductance were measured. Body mass decreased during lactation, but food intake increased. RMR was positively related to the weight of the mammary glands and MEO. We conclude that SusEI is limited at the level of 3.59 × RMR in first-lactating E. miletus. There were no significant effects of litter size on maternal body mass, food intake, thermal conductance, RMR, or litter mass. There were significant effects of fur-shaving on food intake, RMR, and thermal conductance. Fur-shaving increased thermogenic capacity but had no significant effect on reproductive output. These observations provide support for the peripheral limitation hypothesis and argue against the heat dissipation limitation hypothesis.

Food quality and availability are important factors influencing the survival and reproduction of animals. The aim of the present study was to examine the effect of dietary fiber content high-fiber (HF) diet treatment or low-fiber (LF) diet treatment) on energy metabolism, thermogenesis, and leptin concentrations in Chevrier’s field mouse (Apodemus chevrieri (Milne-Edwards, 1868)) (Mammalia: Rodentia: Muridae). Mice on the HF treatment showed a lower body mass compared with LF treatment from day 27 to day 37, and a lower but insignificant body mass to day 71. Dry matter intake (DMI) and gross energy intake (GEI) were greater in HF compared with LF, whereas the digestible energy intake (DEI) was similar for both treatments. Nonshivering thermogenesis (NST) decreased in HF mice, whereas LF mice remained stable; no significant differences were detected in the basal metabolic rate (BMR), uncoupling protein 1 (UCP1) content in brown adipose tissue (BAT), or the levels of serum tri-iodothyronine (T3) and thyroxine (T4) between HF and LF mice. Although there were no differences in body fat content and serum leptin concentrations between HF and LF mice, serum leptin levels were positively correlated with body fat mass. These results support the hypothesis that A. chevrieri can compensate the poor-quality diet physiologically by way of increasing food intake and decreasing thermogenesis.

To investigate the relationship between the energy strategies in response to food restriction and the levels of metabolism in small mammals, body mass, resting metabolic rate (RMR), nonshivering thermogenesis (NST) and cytochrome c oxidase (COX) activity were measured in Apodemus chevrieri that were subjected to different levels of food restriction (FR). The results showed that cold-exposed group had significantly increased RMR and NST, but decreased body mass and survival rate after being restricted to 80% of ad libitum food intake compared with their counterparts maintained at room temperature. A. chevrieri with higher RMR consumed higher food intake than individuals with lower RMR, whereas no differences were observed in body mass and survival rate between two groups after being restricted to 80% of ad libitum food intake. The results suggest that A. chevrieri characterized by higher levels of metabolism are sensitive to periods of FR, providing a support for the “metabolism switch hypothesis”.

This study investigates the energy strategies of a small mammal in response to food shortages as a function of food restriction (FR), metabolic rate and ambient temperature. We subjected tree shrews (Tupaia belangeri) to FR and measured body mass, survival rate, resting metabolic rate (RMR), nonshivering thermogenesis (NST) and cytochromecoxidase (COX) activity of brown adipose tissue (BAT). Cold-exposed animals restricted to 80% ofad libitumfood intake had significantly increased RMR and NST and decreased body mass and survival rates compared with those kept at room temperature on the same FR level. Animals classified has having a high RMR consumed 30.69% more food than those classified as having a low RMR, but showed no differences in body mass or survival when restricted to 80% ofad libitumfood intake. These results indicate that tree shrews, known for their relatively high metabolic rates, are sensitive to periods of FR, which supports the metabolic switch hypothesis. Our findings are also consistent with the prediction that small mammals with food hoarding behaviors, like tree shrews, may have a lower tolerance for food shortages than non-hoarding species.

Chemical communication plays an important role in reproductive and social behaviour of small mammals. The chemical constituents of urine were the main signal resources that can encode sex and social status. The purpose of the present study was to test volatile chemical substances in urine of the kinship and non-kinship Tupaia belangeri, volatile chemicals in urine were performed by the gas chromatography and mass spectrometry (GC-MS), which is speculated that volatile chemicals in urine may had key role in its kin recognition of T. belangeri. The results showed that the components of volatile chemicals in urine were similar between the kinship and non-kinship T. belangeri, which the main components were Alcohols, Alkanes, Esters and Ketones, but the types of each materials were discrepant. “Formic acid, octyl ester” were absence or existence regularly, and “a-Farnesene” and “2,4-Dithiapentane” were found in kinship and non-kinship T. belangeri, which may be signaling substances in the urine. All of the results suggested that volatile chemical substances in urine were different in kinship and non-kinship T. belangeri, indicating that chemical communication based on signals in urine plays an important role in its kin recognition of T. belangeri.

Physiological adjustments are important strategies for small mammals in response to variation in food availability. To determine the physiological mechanisms affected by food restriction and refeeding, tree shrews were restricted to 85% of initial food intake for 4 weeks and refedad libitumfor another 4 weeks. Changes in food intake, body mass, thermogenesis, body composition, mitochondrial cytochromecoxidase activity, uncoupling protein-1 content in brown adipose tissue and serum leptin levels were measured. The results showed that body mass, body fat mass and serum leptin levels significantly decreased in food restricted tree shrews, and increased when the restriction ended, showing a short “compensatory growth” rather than over-weight or obesity compared withad libitumcontrols. Resting metabolic rate, non-shivering thermogenesis, brown adipose tissue mass (mg), and uncoupling protein-1 content decreased significantly in response to food restriction, and returned to the control levels after the animals were refedad libitum, while the brown adipose tissue mass (%) and cytochromecoxidase activity remained stable during food restriction and refeeding. Food intake increased shortly after refeeding, which perhaps contributed to the rapid regaining of body mass. These results suggest thatTupaia belangerican adjust the status of its physiology integratively to cope with the lack of food by means of decreasing body mass, thermogenesis and serum leptin levels. Leptin may act as a starvation signal to predominantly mediate the reduction in body mass and energy expenditure.

Small mammals are usually adapted to cope with changes in food quality and availability. In order to investigate the adaptive strategy of small rodents responding to varying dietary fiber content, in the present study,Apodemus chevrieriindividuals were acclimated to a high-fiber diet for four weeks and then a relatively low-fiber diet for another four weeks. The results show that body mass was relatively stable over the course of acclimation, but dry matter intake, gross energy intake and the mass of the digestive tract increased significantly and digestibility decreased significantly in high-fiber diet mice, while the digestible energy intake was similar for both high-fiber and low-fiber diet mice except for the first week. High-fiber/low-fiber diet mice showed only a significant lower basal metabolic rate and nonshivering thermogenesis compared to low-fiber diet mice on day R1. The high-fiber diet induced a decrease in serum leptin levels and brown adipose tissue mass associated with a reduction in the cytochromecoxidase activity and uncoupling protein 1 content of brown adipose tissue. Body mass, thermogenic capacity, energy intake, serum leptin levels and digestive tract morphology returned to the control levels after 4 weeks of refeeding low-fiber diet. Further, serum leptin levels were positively related to body fat mass and negatively related to food intake. These data indicated that body mass, energy intake, serum leptin levels and organ morphological plasticity were the main strategies by whichA. chevriericopes with variations in dietary fiber content.

Lactation is the most energetically demanding period in small mammals. To investigate the energy balance strategies in lactating Apodemus chevrieri, we compared body mass, energy intake, thermogenic capacity, organ mass and other morphological parameters of the digestive tract of reproductive and non-reproductive females. The results showed that at parturition, mean body mass was 23.89% higher than that of the non-reproductive group. Over the course of the lactation period (from day 0 to day 21), females decreased by 24.30% in body mass and increased by 21.46% in resting metabolic rate (RMR), then all reached the original levels at day 30. Non-shivering thermogenesis (NST) decreased significantly and reached a minimum at peak lactation (on day 21). During lactation, mean NST was 18.55% lower than that of the non-reproductive group. At the same time, lactating A. chevrieri increased their energy intake significantly. Energy intake increased after parturition and reached a maximum at day 21 of lacta...

The hypothalamus and leptin play a key role in the regulation of food intake. The present study investigated the effects of 4 weeks of short- or long-photoperiod on serum leptin levels and food intake in relation to mRNA expression levels of neuropeptide Y, agouti-related protein, pro-opiomelanocortin, and cocaine- and amphetamine-regulated transcript in the hypothalamus of Chevrier’s field mouse (Apodemus chevrieri). There was a significant difference in body fat mass, food intake and neuropeptide Y mRNA expression between the two groups, but serum leptin level, agouti-related protein, pro-opiomelanocortin, and cocaine- and amphetamine-regulated transcript mRNA expression in the hypothalamus were not difference between the two groups. The elevation of neuropeptide Y mRNA regulated neuropeptides in the hypothalamus suggests a physiological role of neuroendocrine factors in food intake during the different photoperiod. We conclude that leptin may be involved in energy balance and body mass regulation.

Shortage of food resources has significant effects on many physiological parameters of animals. The aim of the present study was to examine the relationship between the energetics countermeasures in response to food restriction and the levels of metabolism in Eothenomy miletus. Survival rate, body mass, basal metabolic rate (BMR), nonshivering thermogenesis (NST), body fat mass, serum leptin levels and other physiological parameters were measured. Animals were divided into high-BMR (hBMR) and low-BMR (lBMR) group. The two groups were restricted to 70% of ad libitum food intake for 4 weeks. The data showed that food intake increased by 24.5% in hBMR group than that in lBMR group before the experiment. Body mass, body fat mass, BMR and NST with hBMR or lBMR group significantly decreased after food restricting. Eighty percent of E. miletus survived with hBMR group, but 60% of E. miletus survived with lBMR group. Serum leptin levels were positively correlated with body mass, BMR and NST. The results suggested that E. miletus could apply physiological adjustments to adapt the period of food shortage by reducing energy metabolism, providing the support for the “metabolism switch hypothesis”. E. miletus with hBMR had decreased energy expenditure to maintain the normal physiological function. However, lBMR group could not decrease energy expenditure to meet the stress of available energy resource, which led to body mass decreased and mortality rate increased. Serum leptin levels may be involved in the regulation of energy balance and body mass in E. miletus during the food restriction.

It is generally accepted that dioecious plants occur more frequently in dry and nutrient-poor habitats, suggesting that abiotic stress factors could contribute to evolution of dioecy from hermaphrodite. Therefore, experimental investigations on the responses of subdioecious species, a special sexual system comprising male, female, and hermaphrodite plants, to abiotic stress factors could quantify the contribution of selective pressure on the evolution of dioecy. In this study, we evaluated the physiological responses of different sex morphs of Oxyria sinensis Hemsley, a perennial herb native to the East Himalayas, to drought stress. Male, female, and hermaphrodite plants of O. sinensis were subjected to low, moderate, and high drought stress conditions in a glasshouse. Generally, with increasing water stress, the values of most measured variables slightly decreased, whereas water-use efficiency slightly increased. Furthermore, there were no significant differences in most of the measured parameters among the sex morphs under each drought stress treatment, indicating that O. sinensis might be well-adapted to drought stress conditions as its typical habitat is the dry and hot habitats of xerothermic river valleys. However, nitrogen-use efficiency was significantly higher in male and female plants than in hermaphrodite plants under high drought stress conditions, suggesting that that nitrogen-use efficiency under conditions of drought stress might have contributed to the evolution of dioecy from the hermaphrodite to some degree.