Molecular therapy of obesity and diabetes by a physiological autoregulatory approach

Abstract

Hypothalamic BDNF is a key element in the regulation of energy balance. Here we investigated gene transfer of BDNF in rodent models of obesity and diabetes. BDNF led to marked weight loss and alleviation of obesity-associated insulin resistance. To facilitate clinical translation and ensure BDNF expression dialed down as weight loss progressed, we developed a molecular autoregulatory system using a single rAAV vector harboring two expression cassettes, one constitutively driving BDNF, the other a specific microRNA targeting the therapeutic gene controlled by a promoter (agouti related peptide) responsive to the BDNF-induced physiological changes. Hence, as body weight declined, microRNA expression was activated inhibiting transgene expression and, in contrast to the progressive weight loss associated with a non-regulated approach, led to a plateau once significant weight loss was achieved. This strategy mimics the body's endogenous physiological feedback mechanisms thereby resetting the hypothalamic set point to reverse obesity and metabolic syndrome.Obesity confers significant risk for diabetes, cardiovascular disease, stroke and some cancers 1,2 . Obesity and the related condition, metabolic syndrome (or syndrome X) are increasing rapidly worldwide with significant morbidity and mortality and socioeconomic burden 3 . Life style modifications such as exercise and diet as well as approved drugs have limited efficacy. Bariatric surgery can lead to weight loss at the cost of significant morbidity, underscoring the need for a safer and more effective approach.To identify potential molecular therapeutic candidates, we used an environmental paradigm. In our previous studies, physically and socially more complex housing leads to increased neurogenesis, improved learning and memory and resistance to insults [4][5][6] . Moreover, although fed ad libitum on identical diets, such enriched animals gain less weight than standard housing controls with an improved metabolic profile and insulin sensitivity. To further characterize this phenotype we focused on potential regulators in the hypothalamic arcuate nucleus (Arc), a brain region critical to energy balance. Amongst a number of genes screened, we observed a consistent upregulation in BDNF expression at 2, 4 and 9 weeks of enrichment.BDNF has previously been identified as an important component of the hypothalamic pathway that controls body weight and energy homeostasis 7 . Obese phenotypes are found in BDNF heterozygous mice 8 , a conditional knockout model 9 and focal hypothalamic deletion in adult mice 10 . This mature onset obesity is associated with hyperphagia, hyperleptinemia, hyperinsulinemia and hyperglycemia. Moreover in humans similar symptoms are associated

The synaptoneurosome transcriptome: a model for profiling the emolecular effects of alcohol

“…These results might be due to probe hybridization differences stemming from different splice variants. BDNF is involved in synaptic plasticity, 57,58 obesity 59 and addiction, 60 and potassium channels have been associated with increased sensitivity and tolerance to the sedative effects of ethanol, 61 seizure susceptibility, 62 neonatal familial convulsions and epilepsy 63 and may be important in the withdrawal-induced seizures caused by chronic alcohol consumption in humans.…”

Social overcrowding as a chronic stress model that increases adiposity in mice

“…First-strand cDNA was generated using TaqMan Reverse Transcription Reagent (Applied Biosystems, Roche, Branchburg, NJ, USA) and quantitative PCR was carried out using a LightCycler Sequence Detection System (Roche, Indianapolis, IN, USA) with the Power SYBR Green PCR Master Mix (Applied Biosystems). Specific primers that have been previously validated (Cao et al, 2009; Cao et al, 2010b; Cao et al, 2011; Lin et al, 2011; Liu et al, 2014; McMurphy et al, 2014) were used to detect the following mouse mRNA: Actb (beta-actin), Crh (corticotropin-releasing hormone), Crh1r (corticotropin-releasing hormone receptor 1), Crh2r (corticotropin-releasing hormone receptor 2), Npy (neuropeptide Y), Npy1r (neuropeptide Y receptor Y1), Npy2r (neuropeptide Y receptor Y2), Vgf (nerve growth factor inducible), Avp (arginine vasopressin) , Avpr1a (arginine vasopressin receptor 1A) , Sgk1 (serum/glucocorticoid regulated kinase 1) , Nr3c1 (nuclear receptor subfamily 3, group C, member 1, also known as glucocorticoid receptor), Nr3c2 (nuclear receptor subfamily 3, group C, member 2, also known as mineralocorticoid receptor), Ucp1 (uncoupling protein 1) , Ucp2 (uncoupling protein 2) , Ucp3 (uncoupling protein 3) , Adrb1 (adrenoceptor beta 1) , Adrb2 (adrenoceptor beta 2) , Adrb3 (adrenoceptor beta 3) , Lep (leptin) , Pparg (peroxisome proliferator-activated receptor gamma) , Fasn (fatty acid synthase) , Gpat (glycerol-3-phosphate acyltransferase) , Pomc (proopiomelanocortin) , Agrp (agouti-related peptide) , Mc4r (melanocortin 4 receptor). Specific primers were checked against the NCBI nucleotide database for specificity.…”

Regulation of TRH neurons and energy homeostasis-related signals under stress

“…Support for this role stems from the effects of transferring the Bdnf gene into hypothalami of adult mice, which display decreased body weight and body fat and increased expression of Mc4r, Lepr, Trh, and Crh compared with wild type mice despite having strangely greater than tenfold induction of the orexigenic peptides NPY and AgRP. Under HFD, they gained less weight and little fat compared with normal mice and displayed increased Mc4r, Lepr, Trh, Crh, Cart (Cartpt), and Pomc expression, as well as expression of the metabolic genes of target tissues such as WAT and liver (Cao et al 2009). These results, although not defining the particular hypothalamic nuclei involved, reveals the importance of additional participants such as BDNF in re-setting the response of several hypothalamic-peptidergic systems and their relationship with body weight and adipose tissue.…”