Intergenomic and epistatic interactions control free radical mediated pancreatic ß-cell damage.

Alloxan (AL)-generated Reactive Oxygen Species (ROS) selectively destroy insulin-producing pancreatic ß-cells. A previous genome-wide scan (GWS) using a cohort of 296 F2 hybrids between NOD (AL-sensitive) and ALR (AL-resistant) mice identified linkages contributing to ß-cell susceptibility or resistance to AL-induced diabetes on Chromosomes (Chr) 2, 3, 8, and a single nucleotide polymorphism in mt-Nd2 of the mitochondrial genome (mtDNA). AL treatment of congenic and consomic NOD mouse stocks confirmed resistance linked to both the mtDNA and the Chr 8 locus from ALR [NOD.mtALR.ALR-(D8Mit293-D8Mit137)]. To identify possible epistatic interactions, the GWS analysis was expanded to 678 F2 mice. ALR-derived diabetes-resistance linkages on Chr 8 as well as the mt-Nd2 a allele were confirmed and novel additional linkages on Chr 4, 5, 6, 7, and 13 were identified. Epistasis was observed between the linkages on Chr 8 and 2 and Chr 8 and 6. Furthermore, the mt-Nd2 genotype affected the epistatic interactions between Chr 8 and 2. These results demonstrate that a combination of nuclear-cytoplasmic genome interactions regulates ß-cell sensitivity to ROS-mediated ALD.

Role of TGF-ß in Self-Peptide Regulation of Autoimmunity.

Transforming growth factor (TGF)-ß has been implicated in regulation of the immune system, including autoimmunity. We have found that TGF-ß is readily produced by T cells following immunization with self-peptide epitopes that downregulate autoimmune responses in type 1 diabetes (T1D) prone nonobese diabetic (NOD) mice. These include multiple peptide epitopes derived from the islet ß-cell antigens GAD65 (GAD65 p202-221, GAD65 p217-236), GAD67 (GAD67 p210-229, GAD67 p225-244), IGRP (IGRP p123-145, IGRP p195-214) and insulin B-chain (Ins. B:9-23) that protected NOD mice from T1D. Immunization of NOD mice with the self-MHC class II I-Ag7 ß-chain-derived peptide, I-Aßg7 p54-76 also induced large amounts of TGF-ß and also protected these mice from diabetes development. These results indicate that peptides derived from disease related self-antigens and MHC class II molecules primarily induce TGF-ß producing regulatory Th3 and Tr1-like cells. TGF-ß produced by these cells could enhance the differentiation of induced regulatory iTreg and iTreg17 cells to prevent induction and progression of autoimmune diseases. We therefore suggest that peripheral immune tolerance could be induced and maintained by immunization with self-peptides that induce TGF-ß producing T cells.

The Presence and Preferential Activation of Regulatory T Cells Diminish Adoptive Transfer of Autoimmune Diabetes by Polyclonal Nonobese Diabetic (NOD) T Cell Effectors into NSG versus NOD-scid Mice.

NOD-scid.Il2rg(null) (NSG) mice are currently being used as recipients to screen for pathogenic autoreactive T cells in type 1 diabetes (T1D) patients. We questioned whether the restriction of IL-2R γ-chain (Il-2rγ)-dependent cytokine signaling only to donor cells in NSG recipients differently influenced the activities of transferred diabetogenic T cells when they were introduced as a monoclonal/oligoclonal population versus being part of a polyclonal repertoire. Unexpectedly, a significantly decreased T1D transfer by splenocytes from prediabetic NOD donors was observed in Il-2rγ(null)-NSG versus Il-2rγ-intact standard NOD-scid recipients. In contrast, NOD-derived monoclonal/oligoclonal TCR transgenic ß cell-autoreactive T cells in either the CD8 (AI4, NY8.3) or CD4 (BDC2.5) compartments transferred disease significantly more rapidly to NSG than to NOD-scid recipients. The reduced diabetes transfer efficiency by polyclonal T cells in NSG recipients was associated with enhanced activation of regulatory T cells (Tregs) mediated by NSG myeloid APC. This enhanced suppressor activity was associated with higher levels of Treg GITR expression in the presence of NSG than NOD-scid APC. These collective results indicate NSG recipients might be efficiently employed to test the activity of T1D patient-derived ß cell-autoreactive T cell clones and lines, but, when screening for pathogenic effectors within polyclonal populations, Tregs should be removed from the transfer inoculum to avoid false-negative results.

Genetic Analysis of Substrain Divergence in Non-Obese Diabetic (NOD) Mice.

The non-obese diabetic (NOD) mouse is a polygenic model for type 1 diabetes that is characterized by insulitis, a leukocytic infiltration of the pancreatic islets. During ~35 years since the original inbred strain was developed in Japan, NOD substrains have been established at different laboratories around the world. Although environmental differences among NOD colonies capable of impacting diabetes incidence have been recognized, differences arising from genetic divergence have not been analyzed previously. We use both mouse diversity array and whole-exome capture sequencing platforms to identify genetic differences distinguishing five NOD substrains. We describe 64 single-nucleotide polymorphisms, and two short indels that differ in coding regions of the five NOD substrains. A 100-kb deletion on Chromosome 3 distinguishes NOD/ShiLtJ and NOD/ShiLtDvs from three other substrains, whereas a 111-kb deletion in the Icam2 gene on Chromosome 11 is unique to the NOD/ShiLtDvs genome. The extent of genetic divergence for NOD substrains is compared with similar studies for C57BL6 and BALB/c substrains. As mutations are fixed to homozygosity by continued inbreeding, significant differences in substrain phenotypes are to be expected. These results emphasize the importance of using embryo freezing methods to minimize genetic drift within substrains and of applying appropriate genetic nomenclature to permit substrain recognition when one is used.

Comparison of Two New Mouse Models of Polygenic Type 2 Diabetes at the Jackson Laboratory, NONcNZO10Lt/J and TALLYHO/JngJ.

This review compares two novel polygenic mouse models of type 2 diabetes (T2D), TALLYHO/JngJ and NONcNZO10/LtJ, and contrasts both with the well-known C57BLKS/J-Lepr(db) (db/db) monogenic diabesity model. We posit that the new polygenic models are more representative of the "garden variety" obesity underlying human T2D in terms of their polygenetic rather than monogenic etiology. Moreover, the clinical phenotypes in these new models are less extreme, for example, more moderated development of obesity coupled with less extreme endocrine disturbances. The more progressive development of obesity produces a maturity-onset development of hyperglycemia in contrast to the juvenile-onset diabetes observed in the morbidly obese db/db model. Unlike the leptin receptor-deficient db/db models with central leptin resistance, the new models develop a progressive peripheral leptin resistance and are able to maintain reproductive function. Although the T2D pathophysiology in both TALLYHO/JngJ and NONcNZO10/LtJ is remarkably similar, their genetic etiologies are clearly different, underscoring the genetic heterogeneity underlying T2D in humans.

Genetic and Pharmacologic Models for Type 1 Diabetes.

Type 1 diabetes (T1D) is characterized by a partial or total insufficiency of insulin. The premiere animal model of autoimmune T cell-mediated T1D is the NOD mouse. A dominant negative mutation in the mouse insulin 2 gene (Ins2Akita ) produces a severe insulin deficiency syndrome without autoimmune involvement, as do a variety of transgenes overexpressed in beta cells. Pharmacologically-induced T1D (without autoimmunity) elicted by alloxan or streptozotocin at high doses can generate hyperglycemia in almost any strain of mouse by direct toxicity. Multiple low doses of streptozotocin combine direct beta cell toxicity with local inflammation to elicit T1D in a male sex-specific fashion. A summary of protocols relevant to the management of these different mouse models will be covered in this overview.

Testing the role of P2X7 receptors in the development of type 1 diabetes in nonobese diabetic mice.

Although P2rx7 has been proposed as a type 1 diabetes (T1D) susceptibility gene in NOD mice, its potential pathogenic role has not been directly determined. To test this possibility, we generated a new NOD stock deficient in P2X(7) receptors. T1D development was not altered by P2X(7) ablation. Previous studies found CD38 knockout (KO) NOD mice developed accelerated T1D partly because of a loss of CD4(+) invariant NKT (iNKT) cells and Foxp3(+) regulatory T cells (Tregs). These immunoregulatory T cell populations are highly sensitive to NAD-induced cell death activated by ADP ribosyltransferase-2 (ART2)-mediated ADP ribosylation of P2X(7) receptors. Therefore, we asked whether T1D acceleration was suppressed in a double-KO NOD stock lacking both P2X(7) and CD38 by rescuing CD4(+) iNKT cells and Tregs from NAD-induced cell death. We demonstrated that P2X(7) was required for T1D acceleration induced by CD38 deficiency. The CD38 KO-induced defects in homeostasis of CD4(+) iNKT cells and Tregs were corrected by coablation of P2X(7). T1D acceleration in CD38-deficient NOD mice also requires ART2 expression. If increased ADP ribosylation of P2X(7) in CD38-deficient NOD mice underlies disease acceleration, then a comparable T1D incidence should be induced by coablation of both CD38 and ART2, or CD38 and P2X(7). However, a previously established NOD stock deficient in both CD38 and ART2 expression is T1D resistant. This study demonstrated the presence of a T1D resistance gene closely linked to the ablated Cd38 allele in the previously reported NOD stock also lacking ART2, but not in the newly generated CD38/P2X(7) double-KO line.

mt-Nd2(a) Modifies resistance against autoimmune type 1 diabetes in NOD mice at the level of the pancreatic ß-cell.

OBJECTIVE: To investigate whether a single nucleotide polymorphism (SNP) in the mitochondrial gene for NADH dehydrogenase 2 (mt-Nd2) can modulate susceptibility to type 1 diabetes in NOD mice. RESEARCH DESIGN AND METHODS: NOD/ShiLtJ mice conplastic for the alloxan resistant (ALR)/Lt-derived mt-Nd2(a) allele (NOD.mt(ALR)) were created and compared with standard NOD (carrying the mt-Nd2(c) allele) for susceptibility to spontaneous autoimmune diabetes, or to diabetes elicited by reciprocal adoptive splenic leukocyte transfers, as well as by adoptive transfer of diabetogenic T-cell clones. ß-Cell lines derived from either the NOD (NIT-1) or the NOD.mt(ALR) (NIT-4) were also created to compare their susceptibility to cytolysis by diabetogenic CD8(+) T-cells in vitro. RESULTS: NOD mice differing at this single SNP developed spontaneous or adoptively transferred diabetes at comparable rates and percentages. However, conplastic mice with the mt-Nd2(a) allele exhibited resistance to transfer of diabetes by the CD4(+) T-cell clone BDC 2.5 as well as the CD8(+) AI4 T-cell clones from T-cell receptor transgenic animals. NIT-4 cells with mt-Nd2(a) were also more resistant to AI4-mediated destruction in vitro than NIT-1 cells. CONCLUSIONS: Conplastic introduction into NOD mice of a variant mt-Nd2 allele alone was not sufficient to prevent spontaneous autoimmune diabetes. Subtle nonhematopoietic type 1 diabetes resistance was observed during adoptive transfer experiments with T-cell clones. This study confirms that genetic polymorphisms in mitochondria can modulate ß-cell sensitivity to autoimmune T-cell effectors.

Diet-induced obesity in two C57BL/6 substrains with intact or mutant nicotinamide nucleotide transhydrogenase (Nnt) gene.

The C57BL/6J (B6/J) male mouse represents a standard for diet-induced obesity (DIO) and is unique in expressing a loss-of-function nicotinamide nucleotide transhydrogenase (Nnt) gene. This mutation was associated with a marked reduction in glucose-stimulated insulin secretion from B6/J islets in vitro and moderately impaired glucose clearance in vivo. To assess the contribution of this Nnt mutation, we compared DIO responsiveness of Nnt-mutant B6/J males to Nnt wild-type C57BL/6NJ (B6/NJ) males over a 14-week period of feeding a high-fat (60% of calories) diet. Initial mean body weights at 6 weeks did not distinguish the substrains and both substrains were DIO-sensitive. However, B6/J males outgained the B6/NJ males, with a significant 3 g higher mean body weight at 20 weeks accompanied by significant increases in both lean and fat mass. Mean nonfasting serum glucose over time was also significantly higher in B6/J males, as was impairment of glucose tolerance assessed at 8 and 20 weeks of age. Serum leptin, but not insulin, was significantly higher in B6/J males over time. Potential contributions of the wild-type Nnt gene were demonstrable on a lower fat diet (10% of calories) where a significantly greater weight gain over time by B6/NJ males was correlated with a significantly higher serum insulin. In conclusion, DIO developed in response to 60% fat feeding regardless of Nnt allele status. Contribution of the B6/J-unique Nnt mutation was most evident in response to 10% fat feeding that resulted in reduced serum insulin and weight gain compared to B6/NJ males.

Cdcs1 a major colitis susceptibility locus in mice; subcongenic analysis reveals genetic complexity.

BACKGROUND: The cytokine-deficiency-induced colitis susceptibility (Cdcs)1 locus is a major modifier of murine inflammatory bowel disease (IBD) and was originally identified in experimental crosses of interleukin-10-deficient (Il10(-/-)) mice. Congenic mice, in which this locus was reciprocally transferred between IBD-susceptible C3H/HeJBir-Il10(-/-) and resistant C57BL/6J-Il10(-/-) mice, revealed that this locus likely acts by inducing innate hypo- and adaptive hyperresponsiveness, associated with impaired NF-kappaB responses of macrophages. The aim of the present study was to dissect the complexity of Cdcs1 by further development and characterization of reciprocal Cdcs1 congenic strains and to identify potential candidate genes in the congenic interval. METHODS: In total, 15 reciprocal congenic strains were generated from Il10(-/-) mice of either C3H/HeJBir or C57BL/6J genetic backgrounds by 10 cycles of backcrossing. Colitis activity was monitored by histological grading. Candidate genes were identified by fine mapping of congenic intervals, sequencing, microarray analysis, and a high-throughput real-time reverse-transcription polymerase chain reaction (RT-PCR) approach using bone marrow-derived macrophages. RESULTS: Within the originally identified Cdcs1-interval, 3 independent regions were detected that likely contain susceptibility-determining genetic factors (Cdcs1.1, Cdcs1.2, and Cdcs1.3). Combining results of candidate gene approaches revealed Fcgr1, Cnn3, Larp7, and Alpk1 as highly attractive candidate genes with polymorphisms in coding or regulatory regions and expression differences between susceptible and resistant mouse strains. CONCLUSIONS: Subcongenic analysis of the major susceptibility locus Cdcs1 on mouse chromosome 3 revealed a complex genetic structure. Candidate gene approaches revealed attractive genes within the identified regions.

Animal models of diabetic uropathy.

PURPOSE: Diabetes mellitus is a group of debilitating and costly diseases with multiple serious complications. Lower urinary tract complications or diabetic uropathy are among the most common complications of diabetes mellitus, surpassing widely recognized complications such as neuropathy and nephropathy. Diabetic uropathy develops in individuals with types 1 and 2 diabetes, and little is known about the natural history of these common and troublesome complications. Animal models have the potential to reveal mechanisms and aid in the development of treatment strategies. MATERIALS AND METHODS: We present a review of available animal models of diabetes mellitus relative to their use in the study of diabetic uropathy. RESULTS: Large and small animal models of diabetes mellitus are available. While large animals such as dogs and swine may closely mirror the human disease in size and phenotype, the time between diabetic complication onset and development, and associated husbandry expenditures can make acquiring data on statistically valid sample sizes prohibitively expensive. In contrast, small animal models (rats and mice) have much lower expenditures for a larger number of animals and compressed observation time due to a shorter life span. Also, mice are readily manipulated genetically to facilitate the isolation of the effect of single genes (transgenic and knockout mice). Type 1 diabetes mellitus can be induced chemically with streptozotocin, which is selectively toxic to pancreatic beta cells. Type 2 diabetes mellitus models have been developed by selective breeding for hyperglycemia with or without associated obesity. Diabetic uropathy has been noted in several well characterized, predictable animal models of diabetes mellitus. CONCLUSIONS: Diabetic uropathy, including diabetic bladder dysfunction, has been more frequently studied in small animals with type I diabetes. The recent availability of transgenic models provides a new opportunity for further studies of diabetic uropathy in mouse models of types I and II diabetes mellitus.

Mouse models of diabetic nephropathy.

Diabetic nephropathy is a major cause of ESRD worldwide. Despite its prevalence, a lack of reliable animal models that mimic human disease has delayed the identification of specific factors that cause or predict diabetic nephropathy. The Animal Models of Diabetic Complications Consortium (AMDCC) was created in 2001 by the National Institutes of Health to develop and characterize models of diabetic nephropathy and other complications. This interim report and our online supplement detail the progress made toward that goal, specifically in the development and testing of murine models. Updates are provided on validation criteria for early and advanced diabetic nephropathy, phenotyping methods, the effect of background strain on nephropathy, current best models of diabetic nephropathy, negative models, and views of future directions. AMDCC investigators and other investigators in the field have yet to validate a complete murine model of human diabetic kidney disease. Nonetheless, the critical analysis of existing murine models substantially enhances our understanding of this disease process.

Selecting the "right" mouse model for metabolic syndrome and type 2 diabetes research.

This is not a "Methods" chapter in the traditional sense. Rather, it is an essay designed to help address one of the most frequently asked questions by investigators about to embark on a study requiring an animal model of diabetes - what is the "right" model for the reader's specific research application. Because genetic heterogeneity and the requirement for complex gene-environment interaction characterize the various mouse models of Type 2 diabetes as well as the human disease manifestations, the readers may come to share the author's conclusion that more than one model is required if the investigator is interested in knowing how broadly effective a given compound with putative therapeutic efficacy might be.

Four additional mouse crosses improve the lipid QTL landscape and identify Lipg as a QTL gene.

To identify genes controlling plasma HDL and triglyceride levels, quantitative trait locus (QTL) analysis was performed in one backcross, (NZO/H1Lt x NON/LtJ) x NON/LtJ, and three intercrosses, C57BL/6J x DBA/2J, C57BL/6J x C3H/HeJ, and NZB/B1NJ x NZW/LacJ. HDL concentrations were affected by 25 QTL distributed on most chromosomes (Chrs); those on Chrs 1, 8, 12, and 16 were newly identified, and the remainder were replications of previously identified QTL. Triglyceride concentrations were controlled by nine loci; those on Chrs 1, 2, 3, 7, 16, and 18 were newly identified QTL, and the remainder were replications. Combining mouse crosses with haplotype analysis for the HDL QTL on Chr 18 reduced the list of candidates to six genes. Further expression analysis, sequencing, and quantitative complementation testing of these six genes identified Lipg as the HDL QTL gene on distal Chr 18. The data from these crosses further increase the ability to perform haplotype analyses that can lead to the identification of causal lipid genes.

NOD x 129.H2(g7) backcross delineates 129S1/SvImJ-derived genomic regions modulating type 1 diabetes development in mice.

OBJECTIVE: Introduction of genes targeted in 129/Sv embryonic stem (ES) cells into NOD mice brings about linked genes that may modulate type 1 diabetes. Our objective was to identify 129S1/SvJ non-MHC regions contributing type 1 diabetes resistance or susceptibility in backcross to NOD/LtJ. RESEARCH DESIGN AND METHODS: After congenic transfer of the NOD H2(g7) haplotype onto 129S1/Sv, 310 females were produced by NOD x (NOD x 129.H2(g7))F1 backcross (N2). A genome scan for quantitative trait locus (QTL) affecting clinical diabetes, age of diabetes onset, and insulitis severity was performed using subphenotype characteristics to improve power and resolution for detection of diabetes susceptibility loci. RESULTS: Thirty-six of 310 (11.6%) N2 females developed type 1 diabetes between 14 and 40 weeks. Significant evidence of linkage for only a single previously reported Idd complex locus (Idd10/17/18, chromosome [Chr] 3) was indicated for clinical diabetes. The quantitative traits of insulitis either alone or combined with age at type 1 diabetes onset were significantly linked to known Idd regions on Chr 1 (Idd5 region), Chr 4 (Idd9 region), Chr 8 (Idd22), Chr 11 (Idd4.3), and proximal Chr 17 (Idd16 region). Significant 129S1/Sv resistance contributions were identified on Chr 1, 15 (two loci), and 19, with suggestive evidence for additional novel 129/Sv resistance QTL on Chr 5 and 17 and susceptibility on Chr 2. CONCLUSIONS: The 129S1/SvJ genome harbors collections of both known and potentially novel non-MHC Idd loci. Investigators targeting 129/Sv genes mapping within chromosomal regions reported herein or elsewhere in the genome need to exclude potential contributions from linked Idd loci by generating a NOD.129 control strain expressing the nontargeted allele.

Commonalities of genetic resistance to spontaneous autoimmune and free radical--mediated diabetes.

ALR/Lt, a NOD-related mouse strain, was selected for resistance to alloxan free radical-mediated diabetes (ALD). Despite extensive genomic identity with NOD (>70%), ALR mice display strong resistance to autoimmune type 1 diabetes (T1D) due to both an unusual elevation in systemic antioxidant defenses and a reduction in cellular ROS production that extends to the beta cell level. Reciprocal backcross to NOD previously linked the ALR-derived T1D resistance to Chr. 3, 8, and 17 as well as to the ALR mt-Nd2(a) allele encoded by the mitochondrial genome (mtDNA). To determine whether any of the ALR-derived loci protecting against T1D also protected against ALD, 296 six-week-old F2 mice from reciprocal outcrosses were alloxan-treated and assessed for diabetes onset, and a genome-wide scan (GWS) was conducted. GWS linked mt-Nd2 as well as three nuclear loci with alloxan-induced diabetes. A dominant ALR-derived ALD resistance locus on Chr. 8 colocalized with the ALR-derived T1D resistance locus identified in the previous backcross analysis. In contrast, whereas ALR contributed a novel T1D resistance locus on Chr. 3 marked by Susp, a more proximal ALR-derived region marked by Il-2 contributed ALD susceptibility, not resistance. In addition, a locus was mapped on Chr. 2, where heterozygosity provided heightened susceptibility. Tests for alloxan sensitivity in ALR conplastic mice encoding the NOD mt-Nd2(c) allele and NOD mice congenic for the protective Chr. 8 locus supported our mapping results. Alloxan sensitivity was increased in ALR.mt(NOD) mice, whereas it was decreased by congenic introduction of ALR genome on Chr. 8 into NOD. These data demonstrate both similarities and differences in the genetic control of T1D versus ROS-induced diabetes.

Bone marrow expressing a diabetes resistance MHC class II allele: diabetes deviation by chronic immune stimulation.

The major histocompatibility complex (MHC) of the type 1 diabetes-prone NOD mouse lacks a functional class II H2-Ea gene such that antigen presenting cells (APCs) are I-E null. Transgenic expression of Ea in NOD mice both restores I-E expression and confers complete protection from diabetes progression. Non-myeloablative neonatal transplantation of bone marrow cells from such I-E+ transgenic donors into NOD recipients resulted in low-level but long-term haematopoietic stem cell (HSC) engraftment. Despite low levels of I-E antigen expression in blood (averaging 0.4-3.8% of total MHC class II-positive population), chimeric recipients were protected from overt diabetes, although not insulitis development. Adoptive transfer of diabetes into immunodeficient NOD-Rag recipients that received chimeric splenocytes from primary recipients confirmed the presence of an autoreactive T cell repertoire. The demonstration that purified T cells from these weak chimeras were not tolerant to irradiated transgenic I-E+ splenocytes indicated that I-E+ donor cells provide a constant, low-level immune stimulation capable of up-regulating nominally deficient immunoregulatory networks. This study raises the possibility that cord blood HSCs from infants with high risk HLA haplotypes and a family history of type 1 diabetes might be re-introduced without myoablative treatments following transfection with a single HLA class II allele associated with diabetes resistance.

Gene expression profiling of leukemic cells and primary thymocytes predicts a signature for apoptotic sensitivity to glucocorticoids.

BACKGROUND: Glucocorticoids (GC's) play an integral role in treatment strategies designed to combat various forms of hematological malignancies. GCs also are powerful inhibitors of the immune system, through regulation of appropriate cytokines and by causing apoptosis of immature thymocytes. By activating the glucocorticoid receptor (GR), GCs evoke apoptosis through transcriptional regulation of a complex, interactive gene network over a period of time preceding activation of the apoptotic enzymes. In this study we used microarray technology to determine whether several disparate types of hematologic cells, all sensitive to GC-evoked apoptosis, would identify a common set of regulated genes. We compared gene expression signatures after treatment with two potent synthetic GCs, dexamethasone (Dex) and cortivazol (CVZ) using a panel of hematologic cells. Pediatric CD4+/CD8+ T-cell leukemia was represented by 3 CEM clones: two sensitive, CEM-C7-14 and CEM-C1-6, and one resistant, CEM-C1-15, to Dex. CEM-C1-15 was also tested when rendered GC-sensitive by several treatments. GC-sensitive pediatric B-cell leukemia was represented by the SUP-B15 line and adult B-cell leukemia by RS4;11 cells. Kasumi-1 cells gave an example of the rare Dex-sensitive acute myeloblastic leukemia (AML). To test the generality of the correlations in malignant cell gene sets, we compared with GC effects on mouse non-transformed thymocytes. RESULTS: We identified a set of genes regulated by GCs in all GC-sensitive malignant cells. A portion of these were also regulated in the thymocytes. Because we knew that the highly Dex-resistant CEM-C1-15 cells could be killed by CVZ, we tested these cells with the latter steroid and again found that many of the same genes were now regulated as in the inherently GC-sensitive cells. The same result was obtained when we converted the Dex-resistant clone to Dex-sensitive by treatment with forskolin (FSK), to activate the adenyl cyclase/protein kinase A pathway (PKA). CONCLUSION: Our results have identified small sets of genes that correlate with GC-sensitivity in cells from several hematologic malignancies. Some of these are also regulated in normal mouse thymocytes.

Adipokine and insulin profiles distinguish diabetogenic and non-diabetogenic obesities in mice.

OBJECTIVE: To use longitudinal profiling of plasma adipokines to distinguish diabetogenic vs. non-diabetogenic obesity syndrome in two new mouse models of polygenic obesity. RESEARCH METHODS AND PROCEDURES: Male mice of the NONcNZO5 strain develop a polygenic obesity syndrome uncomplicated by diabetes, whereas NONcNZO10 males develop a comparable polygenic obesity that precipitates type 2 diabetes. A multiplex immunoassay for simultaneous measurement of insulin and a panel of mouse adipokines (leptin, resistin, adiponectin, interleukin-6, tumor necrosis factor alpha, macrophage chemoattractant protein-1, plasminogen activator inhibitor-1) were used to profile longitudinal changes in these strains between 4 and 16 weeks of age that might distinguish the non-diabetogenic vs. diabetogenic obesity (diabesity). RESULTS: Both strains became adipose, with NONcNZO5 males attaining a higher mean body weight with a higher percentage fat content. Weight gain in NONcNZO5 was accompanied by a transient peak in plasma insulin (PI) at 8 weeks followed by a decline into normal range, with normoglycemia maintained throughout. In contrast, NONcNZO10 showed no early PI secretory response because both body weight and plasma glucose increased between 4 and 8 weeks. Only after 12 weeks, with hyperglycemia established, was a delayed PI secretory response observed. Neither plasma leptin nor adiponectin concentrations significantly differentiated the two syndromes over time. However, repeated measures ANOVA showed that NONcNZO10 males maintained significantly higher plasma concentrations of two adipokines, resistin and plasminogen activator inhibitor-1, and the pro-inflammatory cytokine/adipokine macrophage chemoattractant protein-1. DISCUSSION: Longitudinal profiling of PI and adipokines in two new mouse models developing moderate obesity demonstrated that specific marker signatures differentiated a non-diabetogenic obesity from a diabetogenic obesity.