Malgorzata Karbownik-Lewinska

Single‐nucleotide polymorphisms and mRNA expression for melatonin synthesis rate‐limiting enzyme in recurrent depressive disorder

Abstract:  Depressive disorder (DD) is characterised by disturbances in blood melatonin concentration. It is well known that melatonin is involved in the control of circadian rhythms, sleep included. The use of melatonin and its analogues has been found to be effective in depression therapy. Melatonin synthesis is a multistage process, where the last stage is catalysed by acetylserotonin methyltransferase (ASMT), the reported rate‐limiting melatonin synthesis enzyme. Taking into account the significance of genetic factors in depression development, the gene for ASMT may become an interesting focus for studies in patients with recurrent DD. The goal of the study was to evaluate two single‐nucleotide polymorphisms (SNPs) (rs4446909; rs5989681) of the ASMT gene, as well as mRNA expression for ASMT in recurrent DD‐affected patients. We genotyped two polymorphisms in a group of 181 recurrent DD patients and in 149 control subjects. The study was performed using the polymerase chain reaction/restriction fragment length polymorphism method. The distribution of genotypes in both studied SNPs in the ASMT gene differed significantly between DD and healthy subjects. The presence of AA genotype of rs4446909 polymorphism and of GG genotype of rs5989681 polymorphism was associated with lower risk for having recurrent DD. In turn, patients with depression were characterised by reduced mRNA expression for ASMT. In addition, ASMT transcript level in both recurrent DD patients and in healthy subjects depended significantly on genotype distributions in both polymorphisms. In conclusion, our results suggest the ASMT gene as a susceptibility gene for recurrent DD.

The thyroid gland and the process of aging; what is new?

The endocrine system and particular endocrine organs, including the thyroid, undergo important functional changes during aging. The prevalence of thyroid disorders increases with age and numerous morphological and physiological changes of the thyroid gland during the process of aging are well-known. It is to be stressed that the clinical course of thyroid diseases in the elderly differs essentially from that observed in younger individuals, because symptoms are more subtle and are often attributed to normal aging. Subclinical hypo- and hyperthyroidism, as well as thyroid neoplasms, require special attention in elderly subjects. Intriguingly, decreased thyroid function, as well as thyrotropin (TSH) levels – progressively shifting to higher values with age – may contribute to the increased lifespan.This short review focuses on recent findings concerning the alterations in thyroid function during aging, including these which may potentially lead to extended longevity, both in humans and animals.

Expression of Key Regulators of Mitochondrial Biogenesis in Growth Hormone Receptor Knockout (GHRKO) Mice is Enhanced but is Not Further Improved by Other Potential Life-Extending Interventions

Mitochondrial biogenesis is essential for cell viability. Growth hormone receptor knockout (GHRKO), calorie restriction, and surgical visceral fat removal constitute experimental interventions to delay aging and increase life span. We examined the expression of known regulators of mitochondriogenesis: peroxisome proliferator-activated receptor γ co-activator 1α (PGC-1α), adenosine monophosphate (AMP)-activated protein kinase (AMPK), sirtuin-1 (SIRT-1) and sirtuin-3 (SIRT-3), endothelial nitric oxide synthase (eNOS), nuclear respiratory factor-1, mitochondrial transcription factor A (TFAM), and mitofusin-2 (MFN-2) in the skeletal muscles and hearts of control and calorie-restricted female GHRKO mice and in the kidneys of male GHRKOs after visceral fat removal or sham surgery. Expression of PGC-1α in skeletal muscles, AMPK, SIRT-1, SIRT-3, eNOS, and MFN-2 in the heart and PGC-1α, AMPK, SIRT-3, eNOS, and MFN-2 in kidneys was increased in GHRKO mice but was not affected by calorie restriction or visceral fat removal. GHRKO mice have increased expression of key regulators of mitochondriogenesis, which is not improved further by calorie restriction or visceral fat removal.

Single nucleotide polymorphisms and mRNA expression for melatonin MT2 receptor in depression

Polymorphisms (rs 4753426 and rs 794837) and expression of the melatonin MT(2) receptor gene were evaluated in 181 patients with recurrent depressive disorder (rDD) and 149 healthy subjects of Polish origin. We found an increased risk for rDD in patients with the C allele and a decreased risk in patients with the T allele (rs4753426). Patients with the AT heterozygote (rs794837) had an increased mRNA level. The significance of the MT(2) receptor gene and the risk of rDD are suggested.

Oxidative damage to macromolecules in the thyroid - experimental evidence

Whereas oxidative reactions occur in all tissues and organs, the thyroid gland constitutes such an organ, in which oxidative processes are indispensable for thyroid hormone synthesis. It is estimated that huge amount of reactive oxygen species, especially of hydrogen peroxide (H2O2), are produced in the thyroid under physiological conditions, justifying the statement that the thyroid gland is an organ of “oxidative nature”. Apart from H2O2, also other free radicals or reactive species, formed from iodine or tyrosine residues, participate in thyroid hormone synthesis. Under physiological conditions, there is a balance between generation and detoxification of free radicals. Effective protective mechanisms, comprising antioxidative molecules and the process of compartmentalization of potentially toxic molecules, must have been developed in the thyroid to maintain this balance. However, with additional oxidative abuse caused by exogenous or endogenous prooxidants (ionizing radiation being the most spectacular), increased damage to macromolecules occurs, potentially leading to different thyroid diseases, cancer included.

Potassium iodide, but not potassium iodate, as a potential protective agent against oxidative damage to membrane lipids in porcine thyroid

BackgroundFenton reaction (Fe2++H2O2→Fe3++•OH+OH−) is of special significance in the thyroid gland, as both its substrates, i.e. H2O2 and Fe2+, are required for thyroid hormone synthesis. Also iodine, an essential element supplied by the diet, is indispensable for thyroid hormone synthesis. It is well known that iodine affects red-ox balance. One of the most frequently examined oxidative processes is lipid peroxidation (LPO), which results from oxidative damage to membrane lipids. Fenton reaction is used to experimentally induce lipid peroxidation. The aim of the study was to evaluate effects of iodine, used as potassium iodide (KI) or potassium iodate (KIO3), on lipid peroxidation in porcine thyroid homogenates under basal conditions and in the presence of Fenton reaction substrates.MethodsPorcine thyroid homogenates were incubated in the presence of either KI (0.00005 – 500 mM) or KIO3 (0.00005 – 200 mM), without or with addition of FeSO4 (30 μM) + H2O2 (0.5 mM). Concentration of malondialdehyde + 4-hydroxyalkenals (MDA + 4-HDA) was measured spectrophotometrically, as an index of lipid peroxidation.ResultsPotassium iodide, only when used in the highest concentrations (≥50 mM), increased lipid peroxidation in concentration-dependent manner. In the middle range of concentrations (5.0; 10; 25; 50 and 100 mM) KI reduced Fenton reaction-induced lipid peroxidation, with the strongest protective effect observed for the concentration of 25 mM. Potassium iodate increased lipid peroxidation in concentrations ≥2.5 mM. The damaging effect of KIO3 increased gradually from the concentration of 2.5 mM to 10 mM. The strongest damaging effect was observed at the KIO3 concentration of 10 mM, corresponding to physiological iodine concentration in the thyroid. Potassium iodate in concentrations of 5–200 mM enhanced Fenton reaction-induced lipid peroxidation with the strongest damaging effect found again for the concentration of 10 mM.ConclusionsPotassium iodide, used in doses generally recommended in iodide prophylaxis, may prevent oxidative damage to membrane lipids in this gland. Toxic effects of iodide overload may result from its prooxidative action. Potassium iodate does not possess any direct beneficial effects on oxidative damage to membrane lipids in the thyroid, which constitutes an additional argument against its utility in iodine prophylaxis.

High level of oxidized nucleosides in thyroid mitochondrial DNA; damaging effects of Fenton reaction substrates

BackgroundThe mitochondrial DNA (mtDNA) lies in close proximity to the free radical-producing electron transport chain, thus, it is highly prone to oxidative damage. Oxyphilic type of follicular thyroid carcinoma consists of cells filled – almost exclusively – with aberrant mitochondria. In turn, bivalent iron (Fe2+) and hydrogen peroxide (H2O2) are indispensable for thyroid hormone synthesis, therefore being available in physiological conditions presumably at high concentrations. They participate in Fenton reaction (Fe2++H2O2→Fe3++·OH + OH-), resulting in the formation of the most harmful free radical – hydroxyl radical (·OH). The same substrates may be used to experimentally induce oxidative damage to macromolecules. The aim of the study was to evaluate the background level of oxidative damage to mtDNA and the damaging effects of Fenton reaction substrates.MethodsThyroid mtDNA was incubated in the presence of either H2O2 [100, 10, 1.0, 0.5, 0.1, 0.001, 0.00001 mM] or FeSO4 (Fe2+) [300, 150, 30, 15, 3.0, 1.5 μM], or in the presence of those two factors used together, namely, in the presence of Fe2+ [30 μM] plus H2O2 [100, 10, 1.0, 0.5, 0.1, 0.001, 0.00001 mM], or in the presence of H2O2 [0.5 mM] plus Fe2+ [300, 150, 30, 15, 3.0, 1.5 μM]. 8-oxo-7,8-dihydro-2’-deoxyguanosine (8-oxodG) concentration, as the index of DNA damage, was measured by HPLC.ResultsBoth Fenton reaction substrates, used separately, increased 8-oxodG level for the highest H2O2 concentration of 100 mM and in Fe2+ concentration-dependent manner [300, 150, and 30 μM].When Fe2+ and H2O2 were applied together, Fe2+ enhanced H2O2 damaging effect to a higher degree than did H2O2 on Fe2+ effect.ConclusionsThe level of oxidized nucleosides in thyroid mtDNA is relatively high, when compared to nuclear DNA. Both substrates of Fenton reaction, i.e. ferrous ion and hydrogen peroxide, increase oxidative damage to mtDNA, with stronger damaging effect exerted by iron. High level of oxidative damage to mtDNA suggests its possible contribution to malignant transformation of thyroid oncocytic cells, which are known to be especially abundant in mitochondria, the latter characterized by molecular and enzymatic abnormalities.

Key regulators of mitochondrial biogenesis are increased in kidneys of growth hormone receptor knockout (GHRKO) mice

The growth hormone receptor knockout (GHRKO) mice are remarkably long‐lived and highly insulin sensitive. Alterations in mitochondrial biogenesis are associated with aging and various metabolic derangements. We have previously demonstrated increased gene expression of key regulators of mitochondriogenesis in kidneys, hearts and skeletal muscles of GHRKO mice. The aim of the present study was to quantify the protein levels of the following regulators of mitochondriogenesis: peroxisome proliferator‐activated receptor γ co‐activator 1α (PGC‐1α), AMP‐activated protein kinase α (AMPKα), phospho‐AMPKα (p‐AMPKα), sirtuin‐3 (SIRT‐3), endothelial nitric oxide synthase (eNOS), phospho‐eNOS (p‐eNOS), nuclear respiratory factor‐1 (NRF‐1) and mitofusin‐2 (MFN‐2) in skeletal muscles and kidneys of GHRKOs in comparison to normal mice. We also were interested in the effects of calorie restriction (CR) and visceral fat removal (VFR) on these parameters. Both CR and VFR improve insulin sensitivity and can extend life span. Results: The renal levels of PGC‐1α, AMPKα, p‐AMPKα, SIRT‐3, eNOS, p‐eNOS and MFN‐2 were increased in GHRKOs. In the GHRKO skeletal muscles, only MFN‐2 was increased. Levels of the examined proteins were not affected by CR (except for PGC‐1α and p‐eNOS in skeletal muscles) or VFR. Conclusion: GHRKO mice have increased renal protein levels of key regulators of mitochondriogenesis, and this may contribute to increased longevity of these knockouts. Copyright

Decreased thyroid follicle size in dwarf mice may suggest the role of growth hormone signaling in thyroid growth regulation

BackgroundAltered somatotrophic signaling is among the most important potential mechanisms of extended longevity. Ames dwarf (df/df) mice are homozygous for mutation at the Prop-1 gene, leading to a lack of growth hormone (GH), prolactin and thyroid stimulating hormone (TSH). Mice homozygous for targeted disruption of the growth hormone receptor/growth hormone binding protein gene are known as GH receptor knockout (GHRKO) mice or “Laron dwarf”. Both, df/df and GHRKO mice, are characterized by reduced body size, low plasma insulin and insulin-like growth factor-I (IGF-I), remarkably extended longevity, and severe (in df/df mice) or mild (in GHRKO mice) thyroid hypofunction. Recently, by crossing df/df and GHRKO mice, double-mutant Ames dwarf/GHRKO (df/KO) mice were created. Interestingly, these mice are smaller than Ames dwarfs or GHRKOs, and also have reduced insulin and IGF-I levels. The aim of the study was to investigate if and to what extent certain thyroid morphological parameters, such as inner follicular surface area, inner follicular perimeter, as well as the follicular epithelium thickness are changed in the examined dwarf mice.MethodsThis quantification was performed in thyroids collected from df/df, GHRKO and df/KO female mice, at approximately 5–6 months of age. We used a computerized plotting programme that combines a live microscopic image of the slide with an operator-generated overlay.ResultsInner follicular surface area and inner follicular perimeter were decreased in all examined kinds of dwarf mice as compared to normal animals. Furthermore, decreases in these two parameters were more pronounced in df/df and df/KO than in GHRKO mice. Concerning the follicular epithelium thickness, only a tendency towards decrease of this parameter was found in all three kinds of dwarf mice.ConclusionsParameters characterizing thyroid follicle size are decreased in all three examined models of dwarf mice, which may explain decreased thyroid hormone levels in both basal mutants (Ames dwarfs and GHRKOs). df/df mutation seems to predominate over GHRKO genetic intervention concerning their effects on thyroid growth. Beside TSH, also GH signaling seems to constitute a crucial element in the regulation of thyroid growth and, possibly, function.

GH replacement reduces increased lipid peroxidation in GH‐deficient adults

Background  GH replacement improves numerous metabolic abnormalities in GH‐deficient patients; increased lipid peroxidation (LPO) has been observed in GH‐deficient patients; however, it is unknown if LPO is influenced by GH replacement.