Neurosecretory protein GL stimulates food intake, de novo lipogenesis, and onset of obesity
- Graduate School of Integrated Arts and Sciences, Hiroshima University, Japan
- University of California at Berkeley, United States
- Institute for Molecular and Cellular Regulation, Gunma University, Japan
- National Institute for Physiological Sciences, Japan
- School of Life Science, Sokendai (The Graduate University for Advanced Studies), Japan
- Graduate School of Medicine, University of the Ryukyus, Japan
Figures
Figure 1 with 1 supplement
Structure of NPGL and expression of NPGL in rats.
(A) The amino acid structure of NPGL is shown schematically. The bold line between cysteine residues indicates a disulfide bond. (B) Expression levels of the NPGL-precursor mRNA in the entire brain …
Figure 1—figure supplement 1
Amino acid sequences and expression site of NPGL.
(A) Alignment of NPGL-precursor proteins deduced from human, rat, mouse, and chicken cDNA sequences. Black and gray boxes highlight fully conserved and highly conserved amino acids, respectively. …
Figure 2 with 2 supplements
The effects of Npgl overexpression.
The panels show the data obtained by the injection of AAV-based control vector (AAV-CTL) or AAV-based NPGL-precursor gene vector (AAV-NPGL) in high calorie diet (A–D) and normal chow (E–H). (A) …
Figure 2—figure supplement 1
Construction of AAV-based vectors and verification of overexpression.
(A) Schematic structures of AAV-based control vector (AAV-CTL) and AAV-based NPGL-precursor gene vector (AAV-NPGL). Abbreviations: L- and R-ITR: left and right inverted terminal repeat, CMV: …
Figure 2—figure supplement 2
The effects of Npgl overexpression in normal chow.
(A) Changes in daily food intake (n = 6–7). (B) The frequency of various adipocyte sizes measured in 1000 μm2 areas and representative photographs in sections of retroperitoneal WAT (n = 5). Scale …
Figure 3
The effects of Npgl overexpression on O2/CO2 metabolism and lipogenic and lipolytic enzymes.
All panels show data obtained by the injection of AAV-based control vector (AAV-CTL) or AAV-based NPGL-precursor gene vector (AAV-NPGL) in normal chow. (A) O2 consumption (VO2) measured in the …
Figure 4 with 4 supplements
The effects of chronic i.c.v.
infusion of NPGL or antibody against NPGL. All panels show data obtained by the infusion of vehicle (CTL) or NPGL (A–I) in high calorie diet (A–D) and normal chow (E–I), and the control IgG (CTL) or …
Figure 4—figure supplement 1
The effects of chronic i.c.v. infusion of NPGL on lipogenic and lipolytic enzymes.
All panels show data obtained by the infusion of vehicle (CTL) or NPGL in normal chow. (A) mRNA expression levels for lipogenic (Acc, Fas, Scd1, Gpat1, and Adpn) and lipolytic (Cpt1a, Atgl, and Hsl) …
Figure 4—figure supplement 2
The effects of acute i.c.v. injection of NPGL on lipogenic and lipolytic enzymes.
All panels show data obtained after 5 hr of i.c.v. injection of vehicle (CTL) or NPGL in normal chow. mRNA expression levels for lipogenic (Acc, Fas, Scd1, and Gpat1) and lipolytic (Cpt1a, Atgl, and …
Figure 4—figure supplement 3
The effects of chronic i.c.v. infusion of synthetic analogs in normal chow.
(A) Sequences of mature NPGL, and its analogs NPGL-Gly and NPGL(32-80). The two conserved Cys (C) residues, which are involved in the intramolecular disulfide bond formation, are indicated by bold. …
Figure 4—figure supplement 4
The effects of chronic i.c.v. infusion of antibody against NPGL in high calorie diet.
(A) The average intake of a high calorie diet during infusion of control IgG (CTL) or antibody against NPGL (NPGL-Ab) during the light period, dark period, or over 24 hr (n = 8). (B) Body mass (n = 8…
Figure 5 with 1 supplement
Feeding behavior of macronutrient diets during chronic i.c.v. infusion of NPGL.
(A) The calorie intake of macronutrient diets (total, protein, carbohydrate, and fat) by the infusion of vehicle (CTL) or NPGL during the light period, dark period, or over 24 hr (n = 8). (B) Serum …
Figure 5—figure supplement 1
The effects of chronic i.c.v. infusion of NPGL under macronutrient diets.
(A) Changes in calorie intake of a macronutrient diet (total, protein, carbohydrate, and fat) during the light period (n = 8). (B) The changes in calorie intake of macronutrient diets (total, …
Figure 6
The changes of Npgl mRNA by fasting, streptozotocin or insulin and responsibility of NPGL-producing cells to insulin.
(A) The effects of fasting. Npgl mRNA levels in the mediobasal hypothalamus of fasted rats (n = 8). (B) The effects of streptozotocin (STZ) or insulin. Npgl mRNA levels in the mediobasal …
Figure 7
Potential role of NPGL in food intake and fat accumulation.
Downward blue and upward red arrows indicate ‘inhibition’ and ‘stimulation’, respectively and other-colored arrows demonstrate downstream events. (A) During the ordinary feeding state, NPGL …
Tables
Table 1
Blood chemistry during Npgl overexpression.
| A. Blood chemistry during Npgl overexpression under high calorie diet. | ||
|---|---|---|
| AAV-CTL | AAV-NPGL | |
| Glucose (mg/dl) | 144 ± 6.1 | 156 ± 6.2 |
| Free Fatty Acid (mEq/l) | 0.462 ± 0.038 | 0.460 ± 0.033 |
| Triglyceride (mg/dl) | 192 ± 12.7 | 184 ± 13.7 |
| Cholesterol (mg/dl) | 149 ± 9.6 | 127 ± 6.1† |
| Insulin (ng/ml) | 3.56 ± 0.77 | 6.71 ± 0.90* |
| B. Blood chemistry during Npgl overexpression under normal chow. | ||
|---|---|---|
| AAV-CTL | AAV-NPGL | |
| Glucose (mg/dl) | 105 ± 2.6 | 108 ± 2.6 |
| Free Fatty Acid (mEq/l) | 0.703 ± 0.028 | 0.618 ± 0.031† |
| Triglyceride (mg/dl) | 202 ± 13.4 | 253 ± 23.9† |
| Cholesterol (mg/dl) | 80.8 ± 4.0 | 85.4 ± 2.1 |
| Insulin (ng/ml) | 3.52 ± 0.21 | 3.77 ± 0.16 |
| Leptin (ng/ml) | 12.0 ± 0.59 | 18.5 ± 1.60*** |
| Corticosterone (ng/ml) | 511 ± 2.1 | 512 ± 2.1 |
-
†p <0.1, *p<0.05, ***p<0.005.
Table 2
Blood chemistry during chronic i.c.v. infusion of NPGL or the antibody against NPGL.
| A. Blood chemistry during chronic i.c.v. infusion of NPGL under high calorie diet. | ||
|---|---|---|
| CTL | NPGL | |
| Glucose (mg/dl) | 138 ± 5.7 | 135 ± 2.5 |
| Free Fatty Acid (mEq/l) | 0.526 ± 0.043 | 0.56 ± 0.066 |
| Triglyceride (mg/dl) | 101 ± 13.2 | 149 ± 20.9† |
| Cholesterol (mg/dl) | 99.0 ± 5.1 | 120.4 ± 5.3* |
| Insulin (ng/ml) | 2.53 ± 0.45 | 6.03 ± 1.22* |
| Leptin (ng/ml) | 8.76 ± 1.0 | 17.07 ± 1.6*** |
| B. Blood chemistry during chronic i.c.v. infusion of NPGL under normal chow. | ||
|---|---|---|
| CTL | NPGL | |
| Glucose (mg/dl) | 123 ± 7.0 | 128 ± 7.2 |
| Free Fatty Acid (mEq/l) | 0.276 ± 0.017 | 0.289 ± 0.017 |
| Triglyceride (mg/dl) | 73.0 ± 4.5 | 101.7 ± 11.6† |
| Cholesterol (mg/dl) | 66.5 ± 2.1 | 67.5 ± 3.4 |
| Insulin (ng/ml) | 4.51 ± 0.26 | 4.87 ± 0.33 |
| Leptin (ng/ml) | 8.84 ± 1.4 | 7.93 ± 0.70 |
| Corticosterone (ng/ml) | 523 ± 3.6 | 519 ± 1.4 |
| C. Blood chemistry during chronic i.c.v. infusion of the antibody against NPGL under high calorie diet. | ||
|---|---|---|
| CTL | NPGL-Ab | |
| Glucose (mg/dl) | 134 ± 2.38 | 137 ± 1.84 |
| Free Fatty Acid (mEq/l) | 0.529 ± 0.078 | 0.479 ± 0.048 |
| Triglyceride (mg/dl) | 129 ± 14.6 | 118 ± 18.8 |
| Cholesterol (mg/dl) | 111.9 ± 3.7 | 98.5 ± 4.7* |
| Insulin (ng/ml) | 2.23 ± 0.29 | 2.53 ± 0.33 |
| Leptin (ng/ml) | 9.14 ± 0.85 | 7.24 ± 0.51† |
| D. Blood chemistry during chronic i.c.v. infusion of NPGL under macronutrient diet. | ||
|---|---|---|
| CTL | NPGL | |
| Glucose (mg/dl) | 116 ± 4.4 | 124 ± 4.4 |
| Free Fatty Acid (mEq/l) | 0.359 ± 0.034 | 0.416 ± 0.042 |
| Triglyceride (mg/dl) | 74.4 ± 6.4 | 97.7 ± 7.8* |
| Cholesterol (mg/dl) | 99.7 ± 3.5 | 90.8 ± 5.5 |
| Insulin (ng/ml) | 1.94 ± 0.16 | 3.05 ± 0.68 |
| Leptin (ng/ml) | 3.92 ± 0.32 | 6.24 ± 0.33*** |
-
†p <0.1, *p<0.05, ***p<0.005.
Table 3
Sequences of oligonucleotide primers for real-time RT-PCR.
| A. Sequences of oligonucleotide primers for real-time RT-PCR in human tissues. | |||
|---|---|---|---|
| Gene | Forward primer | Reverse primer | Accession no. |
| NPGL | GGAACCATGGCTTAGGAAGG | CCTTAGGAGCTGAGAATATGTA | NM_001102659.1 |
| ACTB | GGCACCACACCTTCTACAAT | AGGTCTCAAACATGATCTGG | NM_001101.3 |
| B. Sequences of oligonucleotide primers for real-time RT-PCR in rat tissues. | |||
|---|---|---|---|
| Gene | Forward primer | Reverse primer | Accession no. |
| Npgl | GGAACCATGGCTTAGGAAGG | TCTAAGGAGCTGAGAATATGCA | LC003309 |
| Acc | GAGGTGGATCAGAGATTTCA | TTCAGCTCTAACTGGAAAGC | NM_022193.1 |
| Fas | AGGATGTCAACAAGCCCAAG | ACAGAGGAGAAGGCCACAAA | NM_017332.1 |
| Scd1 | TGAAAGCTGAGAAGCTGGTG | CAGTGTGGGCAGGATGAAG | NM_139192.2 |
| Gpat1 | CAGCGTGATTGCTACCTGAA | CGGAAGGTGTGGACAAAGAT | NM_017274.1 |
| Adpn | GGGCTACGCTATGTCTGAGC | GAGACTGCACACGAAGGTGA | NM_001282324.1 |
| Cpt1a | GGATGGCATGTGGGTAAAAG | TACTGACACAGGCAGCCAAA | NM_031559.2 |
| Atgl | GCTGCAAGTGGGTTTTTGAT | GTGAACGGTAAGGCACAGGT | NM_001108509.2 |
| Hsl | GAGACGGAGGACCATTTTGA | CGGAGGTCTCTGAGGAACAG | NM_012859.1 |
| Srebp1 | TCACAGATCCAGCAGGTCCCC | GGTCCCTCCACTCACCAGGGT | NM_001276707.1 |
| Lpl | TCTCCTGATGATGCGGATTT | CAACATGCCCTACTGGTTTC | NM_012598.2 |
| Cd36 | GAGGTCCTTACACATACAGAGTTCGTT | ACAGACAGTGAAGGCTCAAAGATG | NM_031561.2 |
| Fatp1 | GCGGCGTTCGGTGTGTAC | GCACGCGGATCAGAACAGA | NM_053580.2 |
| Slc2a2 | GACATCGGTGTGATCAATGC | TGTCGTATGTGCTGGTGTGA | NM_012879 |
| Slc2a4 | CCTCCAGGATGAAGGAAACA | GGGAGAAAAGCCCATCTAGG | NM_012751 |
| Gapdh | CGGCAAGTTCAACGGCACAG | ACTCCACGACATACTCAGCAC | NM_017008.4 |
| Gk | TTGAGACCCGTTTCGTGTCA | AGGGTCGAAGCCCCAGAGT | NM_001270850.1 |
| L-Pk | TGATGATTGGACGCTGCAA | GAGTTGGTCGAGCCTTAGTGATC | NM_012624.3 |
| Npy | TATCCCTGCTCGTGTT | GATTGATGTAGTGTCGCAGA | NM_012614.2 |
| Agrp | GCAGACCGAGCAGAAGATGT | GACTCGTGCAGCCTTACACA | NM_033650.1 |
| Pomc | TAAGAGAGGCCACTGAACAT | GTCTATGGAGGTCTGAAGCA | NM_139326.2 |
| Actb | GGCACCACACTTTCTACAAT | AGGTCTCAAACATGATCTGG | NM_031144.3 |
| Rps18 | AAGTTTCAGCACATCCTGCGAGTA | TTGGTGAGGTCAATGTCTGCTTTC | NM_213557.1 |
| C. Sequences of oligonucleotide primers for real-time RT-PCR in mouse tissues. | |||
|---|---|---|---|
| Gene | Forward primer | Reverse primer | Accession no. |
| Npgl | GGAACCATGGCTTAGGAAGG | TCTAAGGAGCTGAGAATATGCA | LC088498 |
| Actb | GGCACCACACCTTCTACAAT | AGGTCTCAAACATGATCTGG | NM_007393.4 |
Additional files
-
Supplementary file 1
The detailed statistical analysis of figures.
The table shows the statistics test, number of n, comparison, p value, and degree of freedom and F/t/z/ETC value in each figure panel.
- https://doi.org/10.7554/eLife.28527.021
