Phospholipases D1 and D2 Suppress Appetite and Protect against Overweight

<div><p>Obesity is a major risk factor predisposing to the development of peripheral insulin resistance and type 2 diabetes (T2D). Elevated food intake and/or decreased energy expenditure promotes body weight gain and acquisition of adipose tissue. Number of studies implicated phospholipase D (PLD) enzymes and their product, phosphatidic acid (PA), in regulation of signaling cascades controlling energy intake, energy dissipation and metabolic homeostasis. However, the impact of PLD enzymes on regulation of metabolism has not been directly determined so far. In this study we utilized mice deficient for two major PLD isoforms, PLD1 and PLD2, to assess the impact of these enzymes on regulation of metabolic homeostasis. We showed that mice lacking PLD1 or PLD2 consume more food than corresponding control animals. Moreover, mice deficient for PLD2, but not PLD1, present reduced energy expenditure. In addition, deletion of either of the PLD enzymes resulted in development of elevated body weight and increased adipose tissue content in aged animals. Consistent with the fact that elevated content of adipose tissue predisposes to the development of hyperlipidemia and insulin resistance, characteristic for the pre-diabetic state, we observed that <i>Pld1</i>^<i>-/-</i> and <i>Pld2</i>^<i>-/-</i> mice present elevated free fatty acids (FFA) levels and are insulin as well as glucose intolerant. In conclusion, our data suggest that deficiency of PLD1 or PLD2 activity promotes development of overweight and diabetes.</p></div

Deletion of <i>Pld1</i> or <i>Pld2</i> promotes food intake.

<p>A) 24 hours cumulative food intake of 20-weeks old mice with indicated genotypes. B) Oxygen consumption of mice with indicated genotypes at different time of the day. C) Average oxygen consumption during light and dark phase. D) Carbon dioxide production of mice with indicated genotypes at different time of the day. E) Average carbon dioxide during light and dark phase. F) Voluntary movements of mice with indicated genotypes at different time of the day. G) Average voluntary movements of mice during light and dark phase. H) Respiratory exchange rate of mice with indicated genotypes at different time of the day. I) Average respiratory exchange rate of mice during light and dark phase. Data represented as mean +/- S.E.M., n = 6 males for control (black), n = 4 males for <i>Pld1</i>^<i>-/-</i> (orange), n = 6 males for <i>Pld2</i>^<i>-/-</i> (green). *p<0.05, **p<0.01, ***p<0.001</p

Free fatty acids in circulation are elevated in mice deficient for PLD1 or PLD2.

<p>A) Free fatty acids (FFAs), B) Glycerol, C) Triglycerides in circulation of 18-weeks old mice with indicated genotypes. Data represented as mean +/- S.E.M., n = 9 males for control (black), n = 6 males for <i>Pld1</i>^<i>-/-</i> (orange), n = 8 males for <i>Pld2</i>^<i>-/-</i> (green). *p<0.05, **p<0.01, ***p<0.001</p

Deletion of <i>Pld1</i> or <i>Pld2</i> does not affect satiety response.

<p>A) 24 hours cumulative food intake of 20-weeks old mice with indicated genotypes. B) Food intake at different time-points after overnight fasting of 20-weeks old mice with indicated genotypes. Data represented as mean +/- S.E.M., n = 6 females for control (black), n = 4 females for <i>Pld1</i>^<i>-/-</i> (orange), n = 6 females for <i>Pld2</i>^<i>-/-</i> (green). *p<0.05, **p<0.01, ***p<0.001</p

mRNA expression in hypothalamus of neuropeptides controlling food intake.

<p>Relative expression of mRNA in hypothalamus of <i>Pld1</i>^<i>-/-</i>, <i>Pld2</i>^<i>-/-</i> and control mice. The analyzed targets are presented as those with known orexigenic effect: neuropeptide Y (Npy), neuropeptide Y receptor 1 (Npyr1), Agouti Related Neuropeptide (AgRp), Hypocretin (Hcrt), Galanin (Gal); those with anorexigenic effect: Pro-opiomelanocortin (Pomc), Cocaine-amphetamine-regulated transcript (Cart), Corticotropin-releasing factor (Crf), Neuromedin U (Nmu); and those involved in the metabolism of GABA and glutamate: Glutamate-ammonia ligase (Glul), Glutamate decarboxylase (Gad1), Glutaminase (Gls) and 4-aminobutyrate aminotransferase (Abat). Data represented as mean +/- S.E.M., n = 6 males for control (black), n = 5 males for <i>Pld1</i>^<i>-/-</i> (orange), n = 6 males for <i>Pld2</i>^<i>-/-</i> (green). *p<0.05, **p<0.01, ***p<0.001</p

Deletion of Pld1 or Pld2 promotes insulin resistance and glucose intolerance.

<p>A) Insulin tolerance test in <i>Pld1</i>^<i>-/-</i>, <i>Pld2</i>^<i>-/-</i> and control mice at 16 weeks old. n = 7 for control (black), n = 6 for <i>Pld1</i>^<i>-/-</i> (orange), n = 8 for <i>Pld2</i>^<i>-/-</i> (green). B) Area under the curve for the insulin tolerance test. C) Glucose tolerance test in <i>Pld1</i>^<i>-/-</i>, <i>Pld2</i>^<i>-/-</i> and control mice at 18 weeks old. n = 12 for control (black), n = 10 for <i>Pld1</i>^<i>-/-</i> (orange), n = 12 for <i>Pld2</i>^<i>-/-</i> (green). D) Area under the curve for the glucose tolerance test. E) Insulin levels in circulation of mice with indicated genotypes at 20 weeks old. n = 8 males for control (black), n = 6 males for <i>Pld1</i>^<i>-/-</i> (orange), n = 8 males for <i>Pld2</i>^<i>-/-</i> (green). Data represented as mean +/- S.E.M., *p<0.05, **p<0.01, ***p<0.001</p