Historically, physician-scientists have had dual roles in caring for patients and in performing investigative research that could potentially lead to new diagnostics and therapeutics. Physician-scientists conducted teaching rounds in the hospital, surrounded by eager house staff and medical students, and were often avidly pursued as the most important sources of new knowledge for trainees. But alas, times have changed. Now physician-scientists are rarely seen in the hospital; they are most often spotted at their desks tapping out yet another grant application. Most struggle to find the time to mentor students and clinical trainees, let alone to care for patients in the hospital, even though these interactions are often the motivating forces for scientific creativity.
The gastrointestinal (GI) tract is composed of a diverse set of organs that together receive extracorporeal nutrition and convert it to energy substrates and cellular building blocks. In the process, it must sort through all that we ingest and discriminate what is useable from what is not, and having done that, it discards what is “junk.” To accomplish these many and varied tasks, the GI tract relies on endogenous enteric hormones produced by enteroendocrine cells and the enteric nervous system. In many instances, the mediators of these tasks are small peptides that home to the CNS and accessory gut organs to coordinate oral intake with digestive secretions. As the contents of ingested material can contain harmful agents, the gut is armed with an extensive immune system. A breach of the epithelial barrier of the GI tract can result in local and eventually systemic disease if the gut does not mount an aggressive immune response.
Despite substantial fluctuations in daily food intake, animals maintain a remarkably stable body weight, because overall caloric ingestion and expenditure are exquisitely matched over long periods of time, through the process of energy homeostasis. The brain receives hormonal, neural, and metabolic signals pertaining to body-energy status and, in response to these inputs, coordinates adaptive alterations of energy intake and expenditure. To regulate food consumption, the brain must modulate appetite, and the core of appetite regulation lies in the gut-brain axis. This Review summarizes current knowledge regarding the neuroendocrine regulation of food intake by the gastrointestinal system, focusing on gastric distention, intestinal and pancreatic satiation peptides, and the orexigenic gastric hormone ghrelin. We highlight mechanisms governing nutrient sensing and peptide secretion by enteroendocrine cells, including novel taste-like pathways. The increasingly nuanced understanding of the mechanisms mediating gut-peptide regulation and action provides promising targets for new strategies to combat obesity and diabetes.
The gastrointestinal tract has a crucial role in the control of energy homeostasis through its role in the digestion, absorption, and assimilation of ingested nutrients. Furthermore, signals from the gastrointestinal tract are important regulators of gut motility and satiety, both of which have implications for the long-term control of body weight. Among the specialized cell types in the gastrointestinal mucosa, enteroendocrine cells have important roles in regulating energy intake and glucose homeostasis through their actions on peripheral target organs, including the endocrine pancreas. This article reviews the biological actions of gut hormones regulating glucose homeostasis, with an emphasis on mechanisms of action and the emerging therapeutic roles of gut hormones for the treatment of type 2 diabetes mellitus.
Over the past few decades, corticotropin-releasing factor (CRF) signaling pathways have been shown to be the main coordinators of the endocrine, behavioral, and immune responses to stress. Emerging evidence also links the activation of CRF receptors type 1 and type 2 with stress-related alterations of gut motor function. Here, we review the role of CRF receptors in both the brain and the gut as part of key mechanisms through which various stressors impact propulsive activity of the gastrointestinal system. We also examine how these mechanisms translate into the development of new approaches for irritable bowel syndrome, a multifactorial disorder for which stress has been implicated in the pathophysiology.
Celiac disease is characterized by small-intestinal mucosal injury and nutrient malabsorption in genetically susceptible individuals in response to the dietary ingestion of wheat gluten and similar proteins in barley and rye. Disease pathogenesis involves interactions among environmental, genetic, and immunological factors. Although celiac disease is predicted by screening studies to affect approximately 1% of the population of the United States and is seen both in children and in adults, 10%–15% or fewer of these individuals have been diagnosed and treated. This article focuses on the role of adaptive and innate immune mechanisms in the pathogenesis of celiac disease and how current concepts of immunopathogenesis might provide alternative approaches for treating celiac disease.
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.
The association between chronic inflammation and cancer is now well established. This association has recently received renewed interest with the recognition that microbial pathogens can be responsible for the chronic inflammation observed in many cancers, particularly those originating in the gastrointestinal system. A prime example is Helicobacter pylori, which infects 50% of the world’s population and is now known to be responsible for inducing chronic gastric inflammation that progresses to atrophy, metaplasia, dysplasia, and gastric cancer. This Review provides an overview of recent progress in elucidating the bacterial properties responsible for colonization of the stomach, persistence in the stomach, and triggering of inflammation, as well as the host factors that have a role in determining whether gastritis progresses to gastric cancer. We also discuss how the increased understanding of the relationship between inflammation and gastric cancer still leaves many questions unanswered regarding recommendations for prevention and treatment.
Ménétrier disease and gastrointestinal stromal tumors (GISTs) are hyperproliferative disorders of the stomach caused by dysregulated receptor tyrosine kinases (RTKs). In Ménétrier disease, overexpression of TGF-α, a ligand for the RTK EGFR, results in selective expansion of surface mucous cells in the body and fundus of the stomach. In GISTs, somatic mutations of the genes encoding the RTK KIT (or PDGFRA in a minority of cases) result in constitutive kinase activity and neoplastic transformation of gut pacemaker cells (interstitial cells of Cajal). On the basis of the involvement of these RTKs in the pathogenesis of these disorders, Ménétrier disease patients have been effectively treated with a blocking monoclonal antibody specific for EGFR and GIST patients with KIT and PDGFRA tyrosine kinase inhibitors.
Increased insulin secretion and expansion of pancreatic β cell mass work together to maintain normal glucose levels when insulin resistance develops. Changes in glucose concentration have long been known to have profound effects upon the rates of insulin secretion and β cell mass, but various other agents can also cause changes, raising questions about which mechanisms are dominant. Evidence favoring a dominant role for glucose is provided by Terauchi et al. in this issue of the JCI (see the related article beginning on page 246). Mice haploinsufficient for β cell glucokinase (Gck) were unable to increase their β cell mass in response to insulin resistance produced by high-fat feeding. Gck is known to be the glucose sensor for glucose metabolism in β cells. The study also provides strong evidence that insulin receptor substrate 2 (Irs2), which is known to have major effects on β cell growth and survival, is a key downstream mediator of the effects of glucose found in this study.
After intestinal injury, both the number and type of intestinal epithelial cells must be restored. Intestinal stem cells, located at the base of the intestinal crypt, repopulate the depleted crypt in a process known as compensatory proliferation. In this issue of the JCI, Brown et al. describe a new mechanism by which this process is regulated (see the related article beginning on page 258). Surprisingly, they find that a subset of stromal cells present within the intestinal tissue and expressing the proliferative factor prostaglandin-endoperoxidase synthase 2 (Ptgs2) is repositioned next to the intestinal stem cell compartment where local production of PGE2 controls injury-induced epithelial cell proliferation.
Drugs known as beta blockers, which antagonize the β-adrenergic receptor (β-AR), are an important component of the treatment regimen for chronic heart failure (HF). However, a significant body of evidence indicates that genetic heterogeneity at the level of the β1-AR may be a factor in explaining the variable responses of HF patients to beta blockade. In this issue of the JCI, Rochais et al. describe how a single amino acid change in β1-AR alters its structural conformation and improves its functional response to carvedilol, a beta blocker currently used in the treatment of HF (see the related article beginning on page 229). This may explain why some HF patients have better responses not only to carvedilol but to certain other beta blockers as well. The data greatly enhance our mechanistic understanding of myocardial adrenergic signaling and support the development of “tailored” or “personalized” medicine, in which specific therapies could be prescribed based on a patient’s genotype.
Macrophages are present as resident cells in adipose tissue, and blood monocytes are recruited in increased numbers to sites of lipid accumulation in atherosclerosis, a modified form of inflammation in the arterial wall. Recent findings reported by 3 separate groups in this issue of the JCI provide evidence for distinct monocyte subsets, differential chemokine receptor usage, and phenotypic modulation of macrophages in murine models of genetic and high-fat diet–induced disease (see the related articles beginning on pages 175, 185, and 195). These studies raise prospects for selective therapeutic targets to ameliorate macrophage hyperinflammatory responses, while sparing host defense and repair mechanisms.
Unraveling the mechanisms controlling remnant lipoprotein clearance is important, as these lipoproteins are highly atherogenic. The most critical molecule in this process is apoE, which mediates high-affinity binding of remnant lipoproteins to members of the LDL receptor (LDLR) family and cell-surface heparan sulfate proteoglycans (HSPGs), which have been shown to play major independent as well as cooperative roles in remnant lipoprotein clearance. While all the players may have been identified, our understanding of how they interact and function together continues to evolve. In this issue of the JCI, MacArthur et al. (see the related article beginning on page 153) demonstrated that HSPGs under normal physiological conditions are critically important in the clearance of remnant lipoproteins, independent of LDLR family members. The complexity of VLDL and chylomicron remnant clearance was exemplified by the studies of Jones et al., also in this issue (see the related article beginning on page 165). Despite defective clearance of LDL in mice with a deficiency in the adaptor protein controlling internalization of the LDLR, called autosomal recessive hypercholesterolemia (ARH), remnant lipoprotein clearance was not grossly abnormal. A likely explanation is that the abnormal LDLRs bind the remnants and then transfer them to another acceptor for internalization. While the studies clearly demonstrate that the LDLR-related protein 1 is not involved and suggest a role for an additional unidentified receptor, it remains a possibility that HSPGs are responsible for remnant uptake by hepatocytes in the presence of defective LDLR internalization.
The forkhead box m1 (Foxm1) transcription factor is essential for initiation of carcinogen-induced liver tumors; however, whether FoxM1 constitutes a therapeutic target for liver cancer treatment remains unknown. In this study, we used diethylnitrosamine/phenobarbital treatment to induce hepatocellular carcinomas (HCCs) in either WT mice or Arf–/–Rosa26-FoxM1b Tg mice, in which forkhead box M1b (FoxM1b) is overexpressed and alternative reading frame (ARF) inhibition of FoxM1 transcriptional activity is eliminated. To pharmacologically reduce FoxM1 activity in HCCs, we subjected these HCC-bearing mice to daily injections of a cell-penetrating ARF26–44 peptide inhibitor of FoxM1 function. After 4 weeks of this treatment, HCC regions displayed reduced tumor cell proliferation and angiogenesis and a significant increase in apoptosis within the HCC region but not in the adjacent normal liver tissue. ARF peptide treatment also induced apoptosis of several distinct human hepatoma cell lines, which correlated with reduced protein levels of the mitotic regulatory genes encoding polo-like kinase 1, aurora B kinase, and survivin, all of which are transcriptional targets of FoxM1 that are highly expressed in cancer cells and function to prevent apoptosis. These studies indicate that ARF peptide treatment is an effective therapeutic approach to limit proliferation and induce apoptosis of liver cancer cells in vivo.
Antiapoptotic B cell leukemia/lymphoma 2 (BCL2) family proteins are expressed in many cancers, but the circumstances under which these proteins are necessary for tumor maintenance are poorly understood. We exploited a novel functional assay that uses BCL2 homology domain 3 (BH3) peptides to predict dependence on antiapoptotic proteins, a strategy we call BH3 profiling. BH3 profiling accurately predicts sensitivity to BCL2 antagonist ABT-737 in primary chronic lymphocytic leukemia (CLL) cells. BH3 profiling also accurately distinguishes myeloid cell leukemia sequence 1 (MCL1) from BCL2 dependence in myeloma cell lines. We show that the special sensitivity of CLL cells to BCL2 antagonism arises from the requirement that BCL2 tonically sequester proapoptotic BIM in CLL. ABT-737 displaced BIM from BCL2’s BH3-binding pocket, allowing BIM to activate BAX, induce mitochondrial permeabilization, and rapidly commit the CLL cell to death. Our experiments demonstrate that BCL2 expression alone does not dictate sensitivity to ABT-737. Instead, BCL2 complexed to BIM is the critical target for ABT-737 in CLL. An important implication is that in cancer, BCL2 may not effectively buffer chemotherapy death signals if it is already sequestering proapoptotic BH3-only proteins. Indeed, activator BH3-only occupation of BCL2 may prime cancer cells for death, offering a potential explanation for the marked chemosensitivity of certain cancers that express abundant BCL2, such as CLL and follicular lymphoma.
T cell–produced cytokines play a pivotal role in the bone loss caused by inflammation, infection, and estrogen deficiency. IFN-γ is a major product of activated T helper cells that can function as a pro- or antiresorptive cytokine, but the reason why IFN-γ has variable effects in bone is unknown. Here we show that IFN-γ blunts osteoclast formation through direct targeting of osteoclast precursors but indirectly stimulates osteoclast formation and promotes bone resorption by stimulating antigen-dependent T cell activation and T cell secretion of the osteoclastogenic factors RANKL and TNF-α. Analysis of the in vivo effects of IFN-γ in 3 mouse models of bone loss — ovariectomy, LPS injection, and inflammation via silencing of TGF-β signaling in T cells — reveals that the net effect of IFN-γ in these conditions is that of stimulating bone resorption and bone loss. In summary, IFN-γ has both direct anti-osteoclastogenic and indirect pro-osteoclastogenic properties in vivo. Under conditions of estrogen deficiency, infection, and inflammation, the net balance of these 2 opposing forces is biased toward bone resorption. Inhibition of IFN-γ signaling may thus represent a novel strategy to simultaneously reduce inflammation and bone loss in common forms of osteoporosis.
Paget disease is the most exaggerated example of abnormal bone remodeling, with the primary cellular abnormality in the osteoclast. Mutations in the p62 (sequestosome 1) gene occur in one-third of patients with familial Paget disease and in a minority of patients with sporadic Paget disease, with the P392L amino acid substitution being the most commonly observed mutation. However, it is unknown how p62P392L mutation contributes to the development of this disease. To determine the effects of p62P392L expression on osteoclasts in vitro and in vivo, we introduced either the p62P392L or WT p62 gene into normal osteoclast precursors and targeted p62P392L expression to the osteoclast lineage in transgenic mice. p62P392L-transduced osteoclast precursors were hyperresponsive to receptor activator of NF-κB ligand (RANKL) and TNF-α and showed increased NF-κB signaling but did not demonstrate increased 1,25-(OH)2D3 responsivity, TAFII-17 expression, or nuclear number per osteoclast. Mice expressing p62P392L developed increased osteoclast numbers and progressive bone loss, but osteoblast numbers were not coordinately increased, as is seen in Paget disease. These results indicate that p62P392L expression on osteoclasts is not sufficient to induce the full pagetic phenotype but suggest that p62 mutations cause a predisposition to the development of Paget disease by increasing the sensitivity of osteoclast precursors to osteoclastogenic cytokines.
The incretin hormones glucagon-like peptide–1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) control glucose homeostasis through well-defined actions on the islet β cell via stimulation of insulin secretion and preservation and expansion of β cell mass. We examined the importance of endogenous incretin receptors for control of glucose homeostasis through analysis of Glp1r–/–, Gipr–/–, and double incretin receptor knockout (DIRKO) mice fed a high-fat (HF) diet. DIRKO mice failed to upregulate levels of plasma insulin, pancreatic insulin mRNA transcripts, and insulin content following several months of HF feeding. Both single incretin receptor knockout and DIRKO mice exhibited resistance to diet-induced obesity, preservation of insulin sensitivity, and increased energy expenditure associated with increased locomotor activity. Moreover, plasma levels of plasminogen activator inhibitor–1 and resistin failed to increase significantly in DIRKO mice after HF feeding, and the GIP receptor agonist [D-Ala2]GIP, but not the GLP-1 receptor agonist exendin-4, increased the levels of plasma resistin in studies of both acute and chronic administration. These findings extend our understanding of how endogenous incretin circuits regulate glucose homeostasis independent of the β cell via control of adipokine secretion and energy expenditure.
We examined the role of hepatic heparan sulfate in triglyceride-rich lipoprotein metabolism by inactivating the biosynthetic gene GlcNAc N-deacetylase/N-sulfotransferase 1 (Ndst1) in hepatocytes using the Cre-loxP system, which resulted in an approximately 50% reduction in sulfation of liver heparan sulfate. Mice were viable and healthy, but they accumulated triglyceride-rich lipoprotein particles containing apoB-100, apoB-48, apoE, and apoCI-IV. Compounding the mutation with LDL receptor deficiency caused enhanced accumulation of both cholesterol- and triglyceride-rich particles compared with mice lacking only LDL receptors, suggesting that heparan sulfate participates in the clearance of cholesterol-rich lipoproteins as well. Mutant mice synthesized VLDL normally but showed reduced plasma clearance of human VLDL and a corresponding reduction in hepatic VLDL uptake. Retinyl ester excursion studies revealed that clearance of intestinally derived lipoproteins also depended on hepatocyte heparan sulfate. These findings show that under normal physiological conditions, hepatic heparan sulfate proteoglycans play a crucial role in the clearance of both intestinally derived and hepatic lipoprotein particles.
Genetic defects in LDL clearance result in severe hypercholesterolemia and premature atherosclerosis. Mutations in the LDL receptor (LDLR) cause familial hypercholesterolemia (FH), the most severe form of genetic hypercholesterolemia. A phenocopy of FH, autosomal recessive hypercholesterolemia (ARH), is due to mutations in an adaptor protein involved in LDLR internalization. Despite comparable reductions in LDL clearance rates, plasma LDL levels are substantially lower in ARH than in FH. To determine the metabolic basis for this difference, we examined the synthesis and catabolism of VLDL in murine models of FH (Ldlr–/–) and ARH (Arh–/–). The hyperlipidemic response to a high-sucrose diet was greatly attenuated in Arh–/– mice compared with Ldlr–/– mice despite similar rates of VLDL secretion. The rate of VLDL clearance was significantly higher in Arh–/– mice than in Ldlr–/– mice, suggesting that LDLR-dependent uptake of VLDL is maintained in the absence of ARH. Consistent with these findings, hepatocytes from Arh–/– mice (but not Ldlr–/– mice) internalized β-migrating VLDL (β-VLDL). These results demonstrate that ARH is not required for LDLR-dependent uptake of VLDL by the liver. The preservation of VLDL remnant clearance attenuates the phenotype of ARH and likely contributes to greater responsiveness to statins in ARH compared with FH.
Adipose tissue macrophages (ATMs) infiltrate adipose tissue during obesity and contribute to insulin resistance. We hypothesized that macrophages migrating to adipose tissue upon high-fat feeding may differ from those that reside there under normal diet conditions. To this end, we found a novel F4/80+CD11c+ population of ATMs in adipose tissue of obese mice that was not seen in lean mice. ATMs from lean mice expressed many genes characteristic of M2 or “alternatively activated” macrophages, including Ym1, arginase 1, and Il10. Diet-induced obesity decreased expression of these genes in ATMs while increasing expression of genes such as those encoding TNF-α and iNOS that are characteristic of M1 or “classically activated” macrophages. Interestingly, ATMs from obese C-C motif chemokine receptor 2–KO (Ccr2-KO) mice express M2 markers at levels similar to those from lean mice. The antiinflammatory cytokine IL-10, which was overexpressed in ATMs from lean mice, protected adipocytes from TNF-α–induced insulin resistance. Thus, diet-induced obesity leads to a shift in the activation state of ATMs from an M2-polarized state in lean animals that may protect adipocytes from inflammation to an M1 proinflammatory state that contributes to insulin resistance.
Monocytes participate critically in atherosclerosis. There are 2 major subsets expressing different chemokine receptor patterns: CCR2+CX3CR1+Ly-6Chi and CCR2–CX3CR1++Ly-6Clo monocytes. Both C-C motif chemokine receptor 2 (CCR2) and C-X3-C motif chemokine receptor 1 (CX3CR1) are linked to progression of atherosclerotic plaques. Here, we analyzed mouse monocyte subsets in apoE-deficient mice and traced their differentiation and chemokine receptor usage as they accumulated within atherosclerotic plaques. Blood monocyte counts were elevated in apoE–/– mice and skewed toward an increased frequency of CCR2+Ly-6Chi monocytes in apoE–/– mice fed a high-fat diet. CCR2+Ly-6Chi monocytes efficiently accumulated in plaques, whereas CCR2–Ly-6Clo monocytes entered less frequently but were more prone to developing into plaque cells expressing the dendritic cell–associated marker CD11c, indicating that phagocyte heterogeneity in plaques is linked to distinct types of entering monocytes. CCR2– monocytes did not rely on CX3CR1 to enter plaques. Instead, they were partially dependent upon CCR5, which they selectively upregulated in apoE–/– mice. By comparison, CCR2+Ly-6Chi monocytes unexpectedly required CX3CR1 in addition to CCR2 and CCR5 to accumulate within plaques. In many other inflammatory settings, these monocytes utilize CCR2, but not CX3CR1, for trafficking. Thus, antagonizing CX3CR1 may be effective therapeutically in ameliorating CCR2+ monocyte recruitment to plaques without impairing their CCR2-dependent responses to inflammation overall.
Macrophage accumulation participates decisively in the development and exacerbation of atherosclerosis. Circulating monocytes, the precursors of macrophages, display heterogeneity in mice and humans, but their relative contribution to atherogenesis remains unknown. We report here that the Ly-6Chi monocyte subset increased dramatically in hypercholesterolemic apoE–deficient mice consuming a high-fat diet, with the number of Ly-6Chi cells doubling in the blood every month. Ly-6Chi monocytes adhered to activated endothelium, infiltrated lesions, and became lesional macrophages. Hypercholesterolemia-associated monocytosis (HAM) developed from increased survival, continued cell proliferation, and impaired Ly-6Chi to Ly-6Clo conversion and subsided upon statin-induced cholesterol reduction. Conversely, the number of Ly-6Clo cells remained unaffected. Thus, we believe that Ly-6Chi monocytes represent a newly recognized component of the inflammatory response in experimental atherosclerosis.
The TGF-β signaling pathway has a complex role in regulating mammary carcinogenesis. Here we demonstrate that the type III TGF-β receptor (TβRIII, or betaglycan), a ubiquitously expressed TGF-β coreceptor, regulated breast cancer progression and metastasis. Most human breast cancers lost TβRIII expression, with loss of heterozygosity of the TGFBR3 gene locus correlating with decreased TβRIII expression. TβRIII expression decreased during breast cancer progression, and low TβRIII levels predicted decreased recurrence-free survival in breast cancer patients. Restoring TβRIII expression in breast cancer cells dramatically inhibited tumor invasiveness in vitro and tumor invasion, angiogenesis, and metastasis in vivo. TβRIII appeared to inhibit tumor invasion by undergoing ectodomain shedding and producing soluble TβRIII, which binds and sequesters TGF-β to decrease TGF-β signaling and reduce breast cancer cell invasion and tumor-induced angiogenesis. Our results indicate that loss of TβRIII through allelic imbalance is a frequent genetic event during human breast cancer development that increases metastatic potential.
The retinoblastoma tumor suppressor (RB) protein is functionally inactivated in the majority of human cancers and is aberrant in one-third of all breast cancers. RB regulates G1/S-phase cell-cycle progression and is a critical mediator of antiproliferative signaling. Here the specific impact of RB deficiency on E2F-regulated gene expression, tumorigenic proliferation, and the response to 2 distinct lines of therapy was investigated in breast cancer cells. RB knockdown resulted in RB/E2F target gene deregulation and accelerated tumorigenic proliferation, thereby demonstrating that even in the context of a complex tumor cell genome, RB status exerts significant control over proliferation. Furthermore, the RB deficiency compromised the short-term cell-cycle inhibition following cisplatin, ionizing radiation, and antiestrogen therapy. In the context of DNA-damaging agents, this bypass resulted in increased sensitivity to these agents in cell culture and xenograft models. In contrast, the bypass of antiestrogen signaling resulted in continued proliferation and xenograft tumor growth in the presence of tamoxifen. These effects of aberrations in RB function were recapitulated by ectopic E2F expression, indicating that control of downstream target genes was an important determinant of the observed responses. Specific analyses of an RB gene expression signature in 60 human patients indicated that deregulation of this pathway was associated with early recurrence following tamoxifen monotherapy. Thus, because the RB pathway is a critical determinant of tumorigenic proliferation and differential therapeutic response, it may represent a critical basis for directing therapy in the treatment of breast cancer.
Antagonists of β-adrenergic receptors (β-ARs) have become a main therapeutic regimen for the treatment of heart failure even though the mechanisms of their beneficial effects are still poorly understood. Here, we used fluorescent resonance energy transfer–based (FRET-based) approaches to directly monitor activation of the β1-AR and downstream signaling. While the commonly used β-AR antagonists metoprolol, bisoprolol, and carvedilol displayed varying degrees of inverse agonism on the Gly389 variant of the receptor (i.e., actively switching off the β1-AR), surprisingly, only carvedilol showed very specific and marked inverse agonist effects on the more frequent Arg389 variant. These specific effects of carvedilol on the Arg389 variant of the β1-AR were also seen for control of beating frequency in rat cardiac myocytes expressing the 2 receptor variants. This FRET sensor permitted direct observation of activation of the β1-AR in living cells in real time. It revealed that β1-AR variants dramatically differ in their responses to diverse beta blockers, with possible consequences for their clinical use.
Markers of oxidative damage have been detected in brain tissue from patients with Alzheimer disease (AD) and other neurodegenerative disorders. These findings implicate oxidative injury in the neurodegenerative process, but whether oxidative stress is a cause or a consequence of neurotoxicity remains unclear. We used a Drosophila model of human tauopathies to investigate the role of oxidative stress in neurodegeneration. Genetic and pharmacological manipulation of antioxidant defense mechanisms significantly modified neurodegeneration in our model, suggesting that oxidative stress plays a causal role in neurotoxicity. We demonstrate that the JNK signaling pathway is activated in our model, which is in agreement with previous findings in AD tissue. Furthermore, we show that the extent of JNK activation correlates with the degree of tau-induced neurodegeneration. Finally, our findings suggest that oxidative stress acts not to promote tau phosphorylation, but to enhance tau-induced cell cycle activation. In summary, our study identifies oxidative stress as a causal factor in tau-induced neurodegeneration in Drosophila.
Glucokinase (Gck) functions as a glucose sensor for insulin secretion, and in mice fed standard chow, haploinsufficiency of β cell–specific Gck (Gck+/–) causes impaired insulin secretion to glucose, although the animals have a normal β cell mass. When fed a high-fat (HF) diet, wild-type mice showed marked β cell hyperplasia, whereas Gck+/– mice demonstrated decreased β cell replication and insufficient β cell hyperplasia despite showing a similar degree of insulin resistance. DNA chip analysis revealed decreased insulin receptor substrate 2 (Irs2) expression in HF diet–fed Gck+/– mouse islets compared with wild-type islets. Western blot analyses confirmed upregulated Irs2 expression in the islets of HF diet–fed wild-type mice compared with those fed standard chow and reduced expression in HF diet–fed Gck+/– mice compared with those of HF diet–fed wild-type mice. HF diet–fed Irs2+/– mice failed to show a sufficient increase in β cell mass, and overexpression of Irs2 in β cells of HF diet–fed Gck+/– mice partially prevented diabetes by increasing β cell mass. These results suggest that Gck and Irs2 are critical requirements for β cell hyperplasia to occur in response to HF diet–induced insulin resistance.
We identified cellular and molecular mechanisms within the stem cell niche that control the activity of colonic epithelial progenitors (ColEPs) during injury. Here, we show that while WT mice maintained ColEP proliferation in the rectum following injury with dextran sodium sulfate, similarly treated Myd88–/– (TLR signaling–deficient) and prostaglandin-endoperoxide synthase 2–/– (Ptgs2–/–) mice exhibited a profound inhibition of epithelial proliferation and cellular organization within rectal crypts. Exogenous addition of 16,16-dimethyl PGE2 (dmPGE2) rescued the effects of this injury in both knockout mouse strains, indicating that Myd88 signaling is upstream of Ptgs2 and PGE2. In WT and Myd88–/– mice, Ptgs2 was expressed in scattered mesenchymal cells. Surprisingly, Ptgs2 expression was not regulated by injury. Rather, in WT mice, the combination of injury and Myd88 signaling led to the repositioning of a subset of the Ptgs2-expressing stromal cells from the mesenchyme surrounding the middle and upper crypts to an area surrounding the crypt base adjacent to ColEPs. These findings demonstrate that Myd88 and prostaglandin signaling pathways interact to preserve epithelial proliferation during injury using what we believe to be a previously undescribed mechanism requiring proper cellular mobilization within the crypt niche.
Copyright © 2014 American Society for Clinical Investigation