Liver receptor homolog 1 contributes to intestinal tumor formation through effects on cell cycle and inflammation

Liver receptor homolog 1 (LRH-1) is an orphan nuclear receptor that synergizes with beta-catenin/T cell factor 4 signaling to stimulate intestinal crypt cell renewal. We evaluated here the impact of haploinsufficiency of LRH-1 on intestinal tumorigenesis by using two independent mouse models of human colon tumorigenesis. Haploinsufficiency of LRH-1 blunts intestinal tumorigenesis in the ApcMin/+ mice, a genetic model of intestinal cancer. Likewise, Lrh-1+/- mice are protected against the formation of aberrant crypt foci in the colon of mice exposed to the carcinogen azoxymethane. LRH-1 gene expression is reduced in tumors that express elevated levels of the proinflammatory cytokine TNF-alpha. Reciprocally, decreased LRH-1 expression in Lrh-1+/- mice attenuates TNF-alpha expression. Compared with normal human colon, expression and subcellular localization of LRH-1 is significantly altered in neoplastic colon. In combination, these data suggest a role of LRH-1 in the initiation of intestinal tumorigenesis both by affecting cell cycle control as well as through its impact on inflammatory pathways.     

Peroxisome proliferator-activated receptor-gamma co-activator-1alpha (PGC-1alpha) gene polymorphisms and their relationship to Type 2 diabetes in Asian Indians.

AIMS: The objective of the present investigation was to examine the relationship of three polymorphisms, Thr394Thr, Gly482Ser and +A2962G, of the peroxisome proliferator activated receptor-gamma co-activator-1 alpha (PGC-1alpha) gene with Type 2 diabetes in Asian Indians. METHODS: The study group comprised 515 Type 2 diabetic and 882 normal glucose tolerant subjects chosen from the Chennai Urban Rural Epidemiology Study, an ongoing population-based study in southern India. The three polymorphisms were genotyped using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Haplotype frequencies were estimated using an expectation-maximization (EM) algorithm. Linkage disequilibrium was estimated from the estimates of haplotypic frequencies. RESULTS: The three polymorphisms studied were not in linkage disequilibrium. With respect to the Thr394Thr polymorphism, 20% of the Type 2 diabetic patients (103/515) had the GA genotype compared with 12% of the normal glucose tolerance (NGT) subjects (108/882) (P = 0.0004). The frequency of the A allele was also higher in Type 2 diabetic subjects (0.11) compared with NGT subjects (0.07) (P = 0.002). Regression analysis revealed the odds ratio for Type 2 diabetes for the susceptible genotype (XA) to be 1.683 (95% confidence intervals: 1.264-2.241, P = 0.0004). Age adjusted glycated haemoglobin (P = 0.003), serum cholesterol (P = 0.001) and low-density lipoprotein (LDL) cholesterol (P = 0.001) levels and systolic blood pressure (P = 0.001) were higher in the NGT subjects with the XA genotype compared with GG genotype. There were no differences in genotype or allelic distribution between the Type 2 diabetic and NGT subjects with respect to the Gly482Ser and +A2962G polymorphisms. CONCLUSIONS: The A allele of Thr394Thr (G --> A) polymorphism of the PGC-1 gene is associated with Type 2 diabetes in Asian Indian subjects and the XA genotype confers 1.6 times higher risk for Type 2 diabetes compared with the GG genotype in this population.     

Mitochondrial cytochrome P450 (CYP) 1B1 is responsible for melatonin‐induced apoptosis in neural cancer cells

Melatonin is an endogenous indoleamine with a wide range of biological functions in the various organisms from bacteria to mammals. Evidence indicates that melatonin facilitates apoptosis in cancer cells and enhances the antitumor activity of chemotherapy in animals and clinical studies. However, the melatonin metabolism and the key metabolic targets in cancer cells still remain unknown. In this study, U118 and SH‐SY5Y tumor cell lines were used to investigate the metabolic pathways of melatonin in cancer cells. Interestingly, the inhibitory effect of melatonin on proliferation in SH‐SY5Y cells is more potent than that in U118 cells. In contrast, this inhibitory effect on the normal cells is absent. The antitumor effects of melatonin are positively associated with its metabolite N‐acetylserotonin (NAS). Unexpectedly, CYP1B1 is, for first time, identified to localize in the mitochondria of tumor cells, and it metabolizes melatonin to form NAS in situ, which subsequently triggers mitochondria‐dependent apoptosis in cancer cells. In normal cells, NAS does not induce apoptosis. A remarkable individual variation on CYP1B1 expression was also detected in human tumor tissue. These findings provide the novel mechanisms regarding the antitumor effects of melatonin in the level of mitochondria. Thus, we hypothesize that CYP1B1 overexpression in mitochondria would significantly enhance the antitumor effects of melatonin. Mitochondrial CYP1B1 can potentially serve as a specific target to modify the therapeutic and biological effects of melatonin on cancer patients.     

Upregulation of peroxisome proliferator-activated receptor gamma coactivator gene (<Emphasis Type="BoldItalic">PGC1A</Emphasis>) during weight loss is related to insulin sensitivity but not to energy expenditure

Aims/hypothesis We investigated whether skeletal muscle peroxisome proliferator-activated receptor gamma coactivator-1 (PGC1A; also known as PPARGC1A) and its target mitofusin-2 (MFN2), as well as carnitine palmitoyltransferase-1 (CPT1; also known as carnitine palmitoyltransferase 1A [liver] [CPT1A]) and uncoupling protein (UCP)3, are involved in the improvement of insulin resistance and/or in the modification of energy expenditure during surgically induced massive weight loss. Materials and methods Seventeen morbidly obese women (mean BMI: 45.9 ± 4 kg/m²) were investigated before, and 3 and 12 months after, Roux-en-Y gastric bypass (RYGB). We evaluated insulin sensitivity by the euglycaemic–hyperinsulinaemic clamp, energy expenditure and substrate oxidation by indirect calorimetry, and muscle mRNA expression by PCR. Results Post-operatively, PGC1A was enhanced at 3 (p = 0.02) and 12 months (p = 0.03) as was MFN2 (p = 0.008 and p = 0.03 at 3 and 12 months respectively), whereas UCP3 was reduced (p = 0.03) at 12 months. CPT1 did not change. The expression of PGC1A and MFN2 were strongly (p < 0.0001) related. Insulin sensitivity, which increased after surgery (p = 0.002 at 3, p = 0.003 at 12 months), was significantly related to PGC1A and MFN2, but only MFN2 showed an independent influence in a multiple regression analysis. Energy expenditure was reduced at 3 months post-operatively (p = 0.001 vs before RYGB), remaining unchanged thereafter until 12 months. CPT1 and UCP3 were not significantly related to the modifications of energy expenditure or of lipid oxidation rate. Conclusions/interpretation Weight loss upregulates PGC1A, which in turn stimulates MFN2 expression. MFN2 expression significantly and independently contributes to the improvement of insulin sensitivity. UCP3 and CPT1 do not seem to influence energy expenditure after RYGB. G. Gastaldi and A. Russell share first authorship.     

Tumor suppressor function of Liver kinase B1 (Lkb1) is linked to regulation of epithelial integrity

Although loss of epithelial integrity is a hallmark of advanced cancer, it remains poorly understood whether genetic alterations corrupting this integrity causally facilitate tumorigenesis. We show that conditional deletion of tumor suppressor gene Lkb1 (Par-4) in the mammary gland compromises epithelial integrity manifested by mislocalization of cell polarity markers, lateralization of tight junctions, deterioration of desmosomes and basement membrane (BM), and hyperbranching of the mammary ductal tree. We identify the desmosomal BM remodelling serine protease Hepsin as a key factor mediating Lkb1 loss-induced structural alterations in mammary epithelium and BM fragmentation. Although loss of Lkb1 alone does not promote mammary tumorigenesis, combination of Lkb1 deficiency with oncogenic c-Myc leads to dramatic acceleration in tumor formation. The results coupling Lkb1 loss-mediated epithelial integrity defects to mislocalization of serine protease Hepsin and to oncogenic synergy with c-Myc imply that Lkb1 loss facilitates oncogenic proliferation by releasing epithelial cells from structural BM boundaries.     

Tumor suppressor down-regulated in renal cell carcinoma 1 (DRR1) is a stress-induced actin bundling factor that modulates synaptic efficacy and cognition

Stress has been identified as a major causal factor for many mental disorders. However, our knowledge about the chain of molecular and cellular events translating stress experience into altered behavior is still rather scant. Here, we have characterized a murine ortholog of the putative tumor suppressor gene DRR1 as a unique stress-induced protein in brain. It binds to actin, promotes bundling and stabilization of actin filaments, and impacts on actin-dependent neurite outgrowth. Endogenous DRR1 localizes to some, but not all, synapses, with preference for the presynaptic region. Hippocampal virus-mediated enhancement of DRR1 expression reduced spine density, diminished the probability of synaptic glutamate release, and altered cognitive performance. DRR1 emerges as a protein to link stress with actin dynamics, which in addition is able to act on synaptic function and cognition.     

Role of receptor activator of nuclear factor-kappa B ligand (RANKL), osteoprotegerin and macrophage protein 1-alpha (MIP-1a) in monoclonal gammopathy of undetermined significance (MGUS)

The aim of this study was to evaluate the role of markers of bone remodelling, and osteoclast activation/function in patients with monoclonal gammopathy of undetermined significance (MGUS). We have measured serum levels of soluble RANKL (sRANKL), osteoprotegerin (OPG), macrophage inflammatory protein-1alpha (MIP-1alpha), markers of bone resorption [N-telopeptide of collagen type-I (NTX), and tartrate-resistant acid phosphatase isoform-5b (TRACP-5b)] and bone formation [bone-alkaline phosphatase (bALP)] in 40 MGUS patients. These parameters were compared with those of 42 newly diagnosed myeloma patients, and 45 healthy, gender- and age-matched controls. MGUS patients had elevated levels of NTX, sRANKL, and sRANKL/OPG ratio compared with controls (P < 0.0001). Furthermore, TRACP-5b, MIP-1alpha and NTX were decreased in patients with MGUS compared with myeloma patients (P < 0.001), while OPG and bALP were increased (P < 0.001). Serum levels of MIP-1alpha, as well as TRACP-5b, and sRANKL/OPG ratio were reduced, while bALP was increased in MGUS patients, even when compared with myeloma patients who had stage I/II disease. These results demonstrate that increased osteoclastogenesis leading to increased bone resorption is present in MGUS but seems to be compensated for by normal bone formation, which is absent in MM. Furthermore MIP-1alpha, bALP, and sRANKL/OPG may be useful tools for distinguishing between cases of MGUS and early myeloma.     

1 alpha(OH) D3 (Alfarol) is effective for the treatment of chronic B cell leukemia: a case report

We reported a case of chronic B-cell leukemia reacted to the administration of 1 alpha (OH)D3 (Alfarol-CHUGAI Pharm. Co.), The patient showed pancytopenia with IgM-kappa type monoclonal protein in the serum. The bone marrow aspiration was failed due to a dry tap, but the biopsied specimen showed a marked infiltration of small sized lymphoid cells with wide cytoplasm. The leukemic cells from peripheral blood showed a morphology of atypical hairy cells, Surface markers of the leukemic cells were IgM, D(kappa)+, CD 19+, CD 20+, CD 21- and HLADR+, The leukemic cells showed no L-tartrate resistant acid phosphatase sensitivity. This case was diagnosed as a chronic B-cell leukemia closely related to a hairy cell leukemia. The treatment with estrogen-chlorambucil compound (Bestrabucil-KUREHA Chem, Co.) or splenic irradiation was not effective. However, after two months' administration of Alfarol the regular blood transfusion was not needed because of increment of the Hb concentration. After eight months of its administration, the bone marrow aspirate showed a marked decrease in the number of the leukemic cells and a restoration of normal hematopoietic cells. This experience suggested that Alfarol in usefull for the treatment of chronic B cell leukemia including hairy cell leukemia and chronic lymphocytic leukemia.     

Cyclin-dependent kinase 1 (Cdk1) is essential for cell division and suppression of DNA re-replication but not for liver regeneration

Cyclin-dependent kinase 1 (Cdk1) is an archetypical kinase and a central regulator that drives cells through G2 phase and mitosis. Knockouts of Cdk2, Cdk3, Cdk4, or Cdk6 have resulted in viable mice, but the in vivo functions of Cdk1 have not been fully explored in mammals. Here we have generated a conditional-knockout mouse model to study the functions of Cdk1 in vivo. Ablation of Cdk1 leads to arrest of embryonic development around the blastocyst stage. Interestingly, liver-specific deletion of Cdk1 is well tolerated, and liver regeneration after partial hepatectomy is not impaired, indicating that regeneration can be driven by cell growth without cell division. The loss of Cdk1 does not affect S phase progression but results in DNA re-replication because of an increase in Cdk2/cyclin A2 activity. Unlike other Cdks, loss of Cdk1 in the liver confers complete resistance against tumorigenesis induced by activated Ras and silencing of p53.     

Advanced glycation endproduct (AGE) receptor 1 is a negative regulator of the inflammatory response to AGE in mesangial cells

Advanced glycation endproducts (AGE) contribute to kidney disease due to diabetes or aging by means of mesangial cell (MC) receptors, such as the receptor for AGE (RAGE), which promote oxidant-stress-dependent NF-kappaB activation and inflammatory gene expression. MC also express scavenger receptors SR-I and SR-II and AGE receptors 1, 2, and 3 (AGE-R1, -R2, and -R3), some of which are linked to AGE turnover. Because AGE-R1 expression is found suppressed in severe diabetic kidney disease, as other receptors increase, we investigated whether his molecule has a protective role against AGE-induced MC injury. A stable murine MC line overexpressing AGE-R1 (R1-MC) was generated, exhibiting a 1.8- to 2.7-fold increase in (125)I-AGE-specific binding, uptake, and degradation, compared with mock-MC. However, AGE-stimulated NF-kappaB activity and mitogen-activated protein kinase (MAPK) (p44/42) phosphorylation were found markedly suppressed in R1-MC. Additionally, AGE-stimulated macrophage chemotaxis protein 1 and RAGE overexpression were abolished in R1-MC. The effect of R1 on RAGE signaling was investigated after overexpressing RAGE in Chinese hamster ovary cells, which lack RAGE. AGE stimulation elicited NF-kappaB and MAPK activities in RAGE-Chinese hamster ovary cells; however, after cotransfection with R1, these responses were suppressed. Also, after silencing endogenous R1 in wild-type MC by R1 small interfering RNA, AGE-mediated MAPK/p44/42 activation exceeded by >2-fold that of mock-MC, consistent with loss of the activation-inhibitory properties of native AGE-R1. AGE-R1, although enhancing AGE removal, is also a distinct receptor in that it suppresses AGE-mediated MC inflammatory injury through negative regulation of RAGE, a previously uncharacterized pathway that may protect renal and other tissue injury due to diabetes and aging.     

Transmembrane delivery of polypeptide hormones bypassing the intrinsic cell surface receptors: a conjugate of insulin with alpha 2-macroglobulin (alpha 2M) recognizing both insulin and alpha 2M receptors and its biological activity in relation to endocytic pathways

125I-labeled insulin has been cross-linked to alpha 2-macroglobulin (alpha 2M) via a disulfide bond. The resulting insulin-alpha 2M conjugate carried 2.2 insulin moieties per mole of alpha 2M and was able to deliver insulin into rat hepatoma cells H35 and HTC. The insulin delivery was mediated predominantly through alpha 2M receptors and 2 h after binding it was found in the lyposomal fractions in the form of conjugate. When the conjugate was applied to rat hepatoma cells it stimulated activity of tyrosine aminotransferase (TAT) with a potency one-half that of native insulin. Hepatoma cells which were treated with conjugate in the presence of bacitracin were also stimulated for TAT activity. Since bacitracin completely inhibited the alpha 2M binding to its receptors, but inhibited conjugate binding by only 80%, this stimulation must have resulted from the remaining binding of conjugate. These results indicate that the insulin-alpha 2M conjugate was biologically active if it bound to insulin receptors, but that the conjugate bound and internalized through alpha 2M receptors did not act as a mediator for TAT activation. Our results using Percoll density gradients indicate a difference in intracellular processing between insulin, alpha 2M and the conjugate. Mechanisms of action of the conjugate are discussed in relation to the receptor-mediated endocytic pathways.     

Abl-interactor-1 (Abi1) has a role in cardiovascular and placental development and is a binding partner of the alpha4 integrin

Dynamic signals linking the actin cytoskeleton and cell adhesion receptors are essential for morphogenesis during development and normal tissue homeostasis. Abi1 is a central regulator of actin polymerization through interactions with multiple protein complexes. However, the in vivo role of Abi1 remains to be defined. The α4 integrin adhesion receptor is associated with enhanced protrusive activity and regulation of directional cell migration. Among integrin subunits, α4 exhibits unique properties in that it predominantly accumulates at the leading edge of migrating cells; however, the pathways that link the actin-regulatory machinery to α4 at the leading edge have remained elusive. We generated Abi1 KO mice and found that loss of Abi1 phenocopies KO of α4. Mice lacking Abi1 or α4 exhibit midgestational lethality with abnormalities in placental and cardiovascular development. Notably, purified Abi1 protein binds directly to the α4 cytoplasmic tail and endogenous Abi1 colocalizes with phosphorylated α4 at the leading edge of spreading cells. Moreover, Abi1-deficient cells expressing α4 have impaired cell spreading, which is rescued by WT Abi1 but not an Abi1 mutant lacking the α4-binding site. These data reveal a direct link between the α4 integrin and actin polymerization and uncover a role for Abi1 in the regulation of morphogenesis in vivo. The Abi1-α4 interaction establishes a mechanistic paradigm for signaling between adhesion events and enhanced actin polymerization at the earliest stages of protrusion.     

Gly482Ser polymorphism of the peroxisome proliferator-activated receptor-gamma coactivator-1 gene might be a risk factor for diabetic retinopathy in Slovene population (Caucasians) with type 2 diabetes and the Pro12Ala polymorphism of the PPARgamma gene is not

BACKGROUND: The peroxisome proliferator-activated receptor-gamma (PPARgamma) gene has been recently associated with type 2 diabetes, obesity and traits depending on VEGF expression (e.g. retinopathy). The PPARgamma gene and its coactivator, the peroxisome proliferator-activated receptor-gamma coactivator-1 (PPARGC1) gene, have been implicated to be involved in glucose uptake and altered lipid oxidation. We therefore hypothesized that the Gly482Ser polymorphism of the PPARGC1 gene and Pro12Ala polymorphism of the PPARgamma gene might confer susceptibility to diabetic retinopathy in type 2 diabetes. The aim of this study was to investigate the association between the Pro12Ala polymorphism in the PPARgamma gene and Gly482Ser polymorphism in the PPARGC1 gene and the development of diabetic retinopathy in the Slovene population (Caucasians) with type 2 diabetes. METHODS: One hundred and sixty subjects with type 2 diabetes and diabetic retinopathy were compared with 101 diabetic subjects without diabetic retinopathy. Chi-square test was used to compare discrete variables, and continuous clinical data were compared by unpaired students t - test. RESULTS: A significantly higher frequency of the AA genotype of the Gly482Ser polymorphism of the PPARGC1 gene was found in the patients with diabetic retinopathy compared to the patients without diabetic retinopathy (14.4% vs 5.9%; p = 0.035), whereas the Pro12Ala polymorphism of the PPARgamma gene failed to yield an association with diabetic retinopathy. CONCLUSIONS: The present study demonstrates that the AA genotype of the Gly482Ser polymorphism in the PPARGC1 gene might be a risk factor for diabetic retinopathy in the Slovene population (Caucasians) with type 2 diabetes (odds ratio 2.7, 95% confidence interval 1.0-6.8), whereas the Pro12Ala polymorphism of the PPARgamma gene failed to confer susceptibility to diabetic retinopathy.