Brief report Functional promoter polymorphism of the neuronal isoform of tryptophan hydroxylase (Tph2) in suicide

The association between suicide and G-703T polymorphism of the tryptophan hydroxylase 2 (TPH2), the ratelimiting enzyme in the biosynthesis of the neurotransmitter serotonin, was studied in a sample of 291 suicide victims and 280 healthy subjects of Croatian origin. No significant differences were found between the groups. Obtained results do not support involvement of the investigated polymorphism in the susceptibility to suicide completion

Bone morphogenetic protein 1.3 inhibition decreases scar formation and supports cardiomyocyte survival after myocardial infarction

Despite the high prevalence of ischemic heart diseases worldwide, no antibody-based treatment currently exists. Starting from the evidence that a specific isoform of the Bone Morphogenetic Protein 1 (BMP1.3) is particularly elevated in both patients and animal models of myocardial infarction, here we assess whether its inhibition by a specific monoclonal antibody reduces cardiac fibrosis. We find that this treatment reduces collagen deposition and cross-linking, paralleled by enhanced cardiomyocyte survival, both in vivo and in primary cultures of cardiac cells. Mechanistically, we show that the anti-BMP1.3 monoclonal antibody inhibits Transforming Growth Factor β pathway, thus reducing myofibroblast activation and inducing cardioprotection through BMP5. Collectively, these data support the therapeutic use of anti-BMP1.3 antibodies to prevent cardiomyocyte apoptosis, reduce collagen deposition and preserve cardiac function after ischemia

Bone biomarkers and hormones in high-5HT and low-5HT rats.

<p>Depicted are relative differences (%): for the overall difference between the high-5HT (H) and low-5HT (L) sublines, P<0.05 is considered significant; for contrasts between sublines at a given age and between different ages within the same subline, P<0.025 is considered significant (n = 5–10 rats/group). H-L indicates a difference between high-5HT and low-5HT animals. 12–2 indicates a difference between 12 months and 2 months old animals.</p

Physiological characteristics of rats from high-serotonin (5HT) and low-5HT sublines.

<p>A. Indicators of 5HT homeostasis shown as “fold difference” between high- and low-5HT animals with 95% confidence intervals. Reference values were (mean±SD): a) for platelet serotonin level (PSL) 0.80±0.08 μg 5HT/mg platelet protein; b) for platelet serotonin uptake (PSU) 0.69±0.07 nmol 5HT/mg platelet protein/min. Rats were 2 months (PSU measurements) and 12 months (gut <i>Mao-A</i>, <i>Tph1</i> and <i>5HTT</i> expression) of age. PSL and gut 5HT turnover data are given for animals of 2 and 12 months of age. B. No difference in 5HT production and storage in the gut was observed between high- and low-5HT rat sublines. 5HT visualized by using immunohistochemistry was documented at 40× magnification and is depicted by black arrows. C-E. Physical characteristics of high-5HT and low-5HT animals (mean±SD). High-5HT animals are represented by black squares, low-5HT animals by open circles. C—body weight; D—femur length; E—hanging time in the string test (mean values from three 60-sec trials separated by 10-min intervals). Relative differences (%) are shown for subline*age interaction contrasts. P-values were adjusted for multiple comparisons (n = 6–15 rats/group). H-L indicates a difference between high-5HT and low-5HT animals. 12–2 indicates a difference between 12 months and 2 months old animals. Mao-A—monoamine oxidase A; Tph1 –tryptophan hydroxylase 1; 5HTT—serotonin transporter</p

Primer sequences used in RT-PCR analysis.

<p>Primer sequences used in RT-PCR analysis.</p

Systemic role of the peripheral 5HT.

<p>Gut synthesized 5HT enters the platelets via the 5HTT. The quantity of 5HT in platelets depends on the 5HTT activity, while the rate of 5HT synthesis in the gut is equal between both rat sublines (≈ sign). Changes in the serum Ca²⁺ level, influenced by PTH from parathyroid glands and by 1,25(OH)₂D₃ from the kidney, impact the platelet 5HTT activity, with a bidirectional effect on PSL (green-red arrow). Elevated 5HT bidirectionally influences the plasma insulin level (green-red arrow) and induces the hyperthrophy of pancreatic β-cells (dashed arrow), leading to type 2 diabetes with an increased plasma glucose, insulin resistance, glucose intolerance, visceral fat volume and decreased muscle strength. In return, plasma insulin level positively correlates with the PSL (+ sign). Increased insulin and 5HT have an additive effect on bone formation (green arrow). Elevated 5HT increases both bone formation and resorption (larger green arrow), thus increasing the bone turnover and resulting in the net bone loss (large red arrow). 5HT—serotonin, 5HTT—serotonin transporter, PSL—plasma serotonin level, PTH—parathyroid hormone.</p

3D model of trabecular bone reconstructed from μCT images for lumbar spine and distal femur in high-5HT and low-5HT rats at 2 and 12 months of age.

<p>A. Spine—μCT images. B-D. Spine—morphometric indices (mean±SD). E. Femur—μCT images. F-H. Femur—morphometric indices (mean±SD). Shown are relative differences (%): H-L (high-5HT vs. low-5HT animals) at different age; 12–2 months for high and low 5HT animals (n = 8–14 rats/group). Depicted are relative differences (%): for the overall difference between the high-5HT and low-5HT sublines, P<0.05 is considered significant; for contrasts between sublines at a given age and between different ages within the same subline, P<0.025 is considered significant.</p

<i>In vivo</i> effects of <i>Tph1</i> inhibition on platelet serotonin levels (PSL) and bone parameters in 12 months old high 5-HT subline.

<p>On day 1 of the experiment, PSL was measured and treatment with LX1032 (25 mg/kg) (n = 7) or vehicle (control, n = 6) was commenced. At the last day of treatment (Day 36), PSL was determined again. Animals were sacrificed 24 h after the last dose and bone volume (BV/TV, %), trabecular spacing (TbSp, mm) and number of trabecules (Tb.N, 1/mm) were determined in the femur and spine using μCT. A. Data are geometric means (±geometric SD) of PSL values on Day 1 and Day 36. A general linear mixed model (treatment, day [random], treatment*day interaction) was fitted to ln(PSL) and differences (expressed as percentages derived from geometric means ratios) were determined: a) in PSL between the two groups on days 1 and 36; b) in PSL between days 36 and 1, within each group; c) in change in PSL from Day 1 to Day 36 (interaction term coefficient) between the two groups. Adjustment for multiple comparisons was by the simulation method. B. Data are means (±SD) by bone parameter by group, separately for the femur and spine. A separate general linear model was fitted to each of the six ln-transformed outcomes. Differences between groups are expressed as percentage differences (derived from geometric means ratios). C. Non-parametric (Kendall's) regression of bone parameters on change in PSL from the start to the end of treatment. Data depict median slope with confidence interval, Kendall's tau coefficient and P-value.</p

<i>In vitro</i> studies on primary rat osteoblasts and osteoclasts.

<p>A. Alkaline phosphatase staining of primary osteoblasts isolated from 5HT sublines. B. TRAP staining of primary osteoclasts isolated from 5HT sublines. C-D. Expression of mRNA for osteoblast (<i>Alph</i>, <i>Ocn</i>) and osteoclast (<i>Trap</i>, <i>Ctsk</i>) differentiation markers and 5HT-related molecules (<i>5HTT</i>, <i>Tph1</i>, <i>5HT</i>-<i>1B</i>, <i>-2A</i> and -<i>2B</i> receptors) in rat primary osteoblast (C) and osteoclast cultures (D) (n = 3–5). E. Levels of 5HT measured in media from primary high-5HT and low-5HT rat osteoblasts and osteoclasts (n = 4). F. Effects of added 5HT, insulin and their combination on <i>Alph</i> and <i>Ocn</i> expression from primary osteoblasts isolated from high-5HT and low-5HT rats (n = 3). G-H. Effects of added 5HT, insulin and LX1032 on <i>Ctsk</i>, <i>Trap</i>, and <i>Tph1</i> mRNA expression (G) (n = 3) and osteoclast number (H) (n = 4–7 wells analyzed per group) in primary osteoclast cultures from control Wistar rats. Expression data (C, D, F, G) are shown as fold difference over a control (indicated by the dashed line) with 95% confidence intervals. Stars (*) at estimates indicate significance vs. control, whereas horizontal lines and associated stars indicate significance in fold difference over control between treatments.</p