69 research outputs found
Uptake and turnover of mono-iodinated thyroid hormone metabolites by PCCL3 thyrocytes
Tyrosine and phenolic ring de-iodination of thyroid hormones (TH) is crucial
for regulating their physiological activity. Furthermore, reactions such as
de-carboxylation to thyronamines (TAM) and de-amination to thyroacetic acids
(TAc) produce TH metabolites (THM) with distinct biological properties. This
needs to be considered when studying effects of TH and THM. The accurate and
precise quantitative analysis of TH and THM in cell culture supernatants and
cell lysates are key procedures required for studying the in vitro metabolism
of TH. We report here the development of a liquid-liquid extraction/isotope
dilution-liquid chromatography-electrospray tandem mass spectrometry (LC-
MS/MS) method for the quantification of 9 thyronines (TN) and 6 TAM in human
hepatocellular carcinoma Hep G2 cell lysate extracts. In addition, we adapted
the method to quantify TH, TAM and TAc, in cell lysates of FBS-depleted rat
thyroid epithelium PCCL3 cells. The methods for both cell lines were validated
by rigorous assessment of linearity, limits of quantification and detection
(LLOQ and LLOD respectively), intra- and inter-day accuracy, precision,
process efficiency (PE), matrix effect (ME) and relative recovery (RE).
Calibration curves covering 11 concentrations (based on 400 μl of lysate) were
linear in the range 0.016–50 nM and 0.010–50 nM for Hep G2 and PCCL3 cells
respectively. The lower limits of quantification were in the range 0.031 to 1
nM. We applied the PCCL3 version of the LC-MS/MS method to the analysis of
lysed cell extracts from PCCL3 cells that had been incubated with
3-iodo-L-thyronine (T1), 3-iodothyronamine (3-T1AM) and 3-iodothyroacetic acid
(3-T1Ac). Over the course of 30 minutes incubation 3-T1AM was de-iodinated to
4-[4-(2-aminoethylphenoxy)]phenol (thyronamine, T0AM) and de-aminated to
3-T1Ac respectively, whilst T1 underwent de-iodination to T0. This data
indicates avid metabolism of these mono-iodinated compounds and the utility of
LC-MS/MS to quantify such cellular metabolism
Alternation between short- and long photoperiod reveals hypothalamic gene regulation linked to seasonal body weight changes in Djungarian hamsters (Phodopus sungorus)
This work was funded by the German Research Foundation (DFG, Emmy-Noether HE6383 to AH) and the British Society for Neuroendocrinology (Research grant to JB). The authors declare that they have no conflicts of interest.Peer reviewedPostprin
Prolonged hypothyroidism severely reduces ovarian follicular reserve in adult rats
Background There is substantial evidence both in humans and in animals that a
prolonged reduction in plasma thyroid hormone concentration leads to
reproductive problems, including disturbed folliculogenesis, impaired
ovulation and fertilization rates, miscarriage and pregnancy complications.
The objective of the present study is to examine the consequences of chronic
hypothyroidism, induced in adulthood, for the size of the ovarian follicle
pool. In order to investigate this, adult female rats were provided either a
control or an iodide deficient diet in combination with perchlorate
supplementation to inhibit iodide uptake by the thyroid. Sixteen weeks later
animals were sacrificed. Blood was collected for hormone analyses and ovaries
were evaluated histologically. Results At the time of sacrifice, plasma
thyroid-stimulating hormone concentrations were 20- to 40-fold increased,
thyroxine concentrations were negligible while tri-iothyronin concentrations
were decreased by 40% in the hypothyroid group, confirming that the animals
were hypothyroid. Primordial, primary and preantral follicle numbers were
significantly lower in the hypothyroid ovaries compared to the euthyroid
controls, while a downward trend in antral follicle and corpora lutea numbers
was observed. Surprisingly the percentage of atretic follicles was not
significantly different between the two groups, suggesting that the reduced
preantral and antral follicle numbers were presumably not the consequence of
increased degeneration of these follicle types in the hypothyroid group.
Plasma anti-Müllerian hormone (AMH) levels showed a significant correlation
with the growing follicle population represented by the total ovarian number
of primary, preantral and antral follicles, suggesting that also under
hypothyroid conditions AMH can serve as a surrogate marker to assess the
growing ovarian follicle population. Conclusions The induction of a chronic
hypothyroid condition in adult female rats negatively affects the ovarian
follicular reserve and the size of the growing follicle population, which may
impact fertility
Sex-specific associations of serum selenium and selenoprotein P with type 2 diabetes mellitus and hypertension in the Berlin Aging Study II
Background: Selenium is essential for expression and proper function of a set of redox active selenoproteins implicated in aging-relevant diseases, e.g. type 2 diabetes mellitus (T2D) and hypertension. However, data in cohorts of older adults, particularly with respect to different Se biomarkers and sex-specific analyses are sparse.
Objective: To assess associations of serum Se and selenoprotein P (SELENOP) concentrations with T2D and hypertension in a cohort of older females and males.
Methods: This study included 1500 participants from the Berlin Aging Study II. Diagnosis of T2D was made in case of antidiabetic medication, self-reported T2D, or laboratory parameters. Diagnosis of hypertension was based on self-report, blood pressure measurement, or anti-hypertensive medication. Se was measured by spectroscopy, and SELENOP by ELISA. Multiple adjusted regression models quantified dose-dependent associations.
Results: Participants had a median(IQR) age of 68 (65,71) years, and 767 (51%) were women. 191 (13%) participants had T2D and 1126 (75%) had hypertension. Se and SELENOP correlated significantly (r = 0.59, p < 0.001), and were elevated in those with self-reported Se supplementation. Serum Se and SELENOP were not associated with T2D in the whole cohort. In men, SELENOP was positively associated with T2D, OR (95%CI) for one mg/L increase in SELENOP was 1.22 (1.00,1.48). Se was non-linearly associated with hypertension, comparing to the lowest quartile (Q1), and participants with higher Se levels (Q3) had a lower OR (95%CI) of 0.66 (0.45,0.96), which was specific for men. SELENOP positively associated with hypertension, and OR (95%CI) per one mg/L increase was 1.15 (1.01,1.32).
Conclusions: The data suggest a sex-specific interrelationship of Se status with T2D and hypertension, with apparent biomarker-specific associations
Dual Effect on Adult-Type Leydig Cell and Sertoli Cell Development
Transient neonatal 6-propyl-2-thiouracil (PTU) induced hypothyroidism affects
Leydig and Sertoli cell numbers in the developing testis, resulting in
increased adult testis size. The hypothyroid condition was thought to be
responsible, an assumption questioned by studies showing that uninterrupted
fetal/postnatal hypothyroidism did not affect adult testis size. Here, we
investigated effects of transient hypothyroidism on Leydig and Sertoli cell
development, employing a perinatal iodide-deficient diet in combination with
sodium perchlorate. This hypothyroidism inducing diet was continued until days
1, 7, 14, or 28 postpartum (pp) respectively, when the rats were switched to a
euthyroid diet and followed up to adulthood. Continuous euthyroid and
hypothyroid, and neonatal PTU-treated rats switched to the euthyroid diet at
28 days pp, were included for comparison. No effects on formation of the
adult-type Leydig cell population or on Sertoli cell proliferation and
differentiation were observed when the diet switched at/or before day 14 pp.
However, when the diet was discontinued at day 28 pp, Leydig cell development
was delayed similarly to what was observed in chronic hypothyroid rats.
Surprisingly, Sertoli cell proliferation was 6- to 8-fold increased 2 days
after the diet switch and remained elevated the next days. In adulthood,
Sertoli cell number per seminiferous tubule cross-section and consequently
testis weight was increased in this group. These observations implicate that
increased adult testis size in transiently hypothyroid rats is not caused by
the hypothyroid condition per se, but originates from augmented Sertoli cell
proliferation as a consequence of rapid normalization of thyroid hormone
concentrations
The effect of high dose isoflavone supplementation on serum reverse T3 in euthyroid men with Type 2 Diabetes and post-menopausal women
Background: The health benefits of soy are widely reported but there are queries on the effect of soy isoflavones on thyroid function and the underlying mechanism of action.Materials and Methods: We examined the effect of soy isoflavones on reverse tri-iodothyronine (or 3,3′,5′-tri-iodothyronine; rT3) in two studies comprising 400 patients: 200 men (study 1; 3 months) and 200 post-menopausal women (study 2; 6 months) who were randomized to consume 15 g soy protein with 66 mg of isoflavones (SPI) daily, or 15 g soy protein alone without isoflavones (SP) daily.Results: SPI supplementation increased rT3 serum concentration in both men 0.41 (0.12) vs. 0.45 (0.14) nmol/L and women 0.33 (0.12) vs. 0.37 (0.09) nmol/L at 3 months compared to SP that was not seen at 6 months. Thyroid stimulating hormone (TSH) serum concentrations increased while free thyroxine (fT4) concentrations decreased with 3 months of SPI compared to SP supplementation for both men and women. rT3 correlated with TSH in both studies (p = 0.03) but not with either fT3 or fT4. fT3 levels did not differ between the SPI and SP preparations.Conclusion: Soy isoflavones transiently increased rT3 levels within 3 months though reverted to baseline at 6 months. The mechanism for this would be either rT3 degrading deiodinase 1 and/or deiodinase 2 activities are transiently inhibited at 3 months, or inhibition of deiodinase 3, which generates rT3 from T4 is induced at 6 months. These changes were mirrored in the TSH concentrations, suggesting that short-term high dose isoflavone transiently impairs thyroid function in the first 3 months and may impact on general health during this period
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3,5-T2-A Janus-Faced Thyroid Hormone Metabolite Exerts Both Canonical T3-Mimetic Endocrine and Intracrine Hepatic Action.
Over the last decades, thyroid hormone metabolites (THMs) received marked attention as it has been demonstrated that they are bioactive compounds. Their concentrations were determined by immunoassay or mass-spectrometry methods. Among those metabolites, 3,5-diiodothyronine (3,5-T2), occurs at low nanomolar concentrations in human serum, but might reach tissue concentrations similar to those of T4 and T3, at least based on data from rodent models. However, the immunoassay-based measurements in human sera revealed remarkable variations depending on antibodies used in the assays and thus need to be interpreted with caution. In clinical experimental approaches in euthyroid volunteers and hypothyroid patients using the immunoassay as the analytical tool no evidence of formation of 3,5-T2 from its putative precursors T4 or T3 was found, nor was any support found for the assumption that 3,5-T2 might represent a direct precursor for serum 3-T1-AM generated by combined deiodination and decarboxylation from 3,5-T2, as previously documented for mouse intestinal mucosa. We hypothesized that lowered endogenous production of 3,5-T2 in patients requiring T4 replacement therapy after thyroidectomy or for treatment of autoimmune thyroid disease, compared to production of 3,5-T2 in individuals with intact thyroid glands might contribute to the discontent seen in a subset of patients with this therapeutic regimen. So far, our observations do not support this assumption. However, the unexpected association between high serum 3,5-T2 and elevated urinary concentrations of metabolites related to coffee consumption requires further studies for an explanation. Elevated 3,5-T2 serum concentrations were found in several situations including impaired renal function, chronic dialysis, sepsis, non-survival in the ICU as well as post-operative atrial fibrillation (POAF) in studies using a monoclonal antibody-based chemoluminescence immunoassay. Pilot analysis of human sera using LC-linear-ion-trap-mass-spectrometry yielded 3,5-T2 concentrations below the limit of quantification in the majority of cases, thus the divergent results of both methods need to be reconciliated by further studies. Although positive anti-steatotic effects have been observed in rodent models, use of 3,5-T2 as a muscle anabolic, slimming or fitness drug, easily obtained without medical prescription, must be advised against, considering its potency in suppressing the HPT axis and causing adverse cardiac side effects. 3,5-T2 escapes regular detection by commercially available clinical routine assays used for thyroid function tests, which may be seriously disrupted in individuals self-administering 3,5-T2 obtained over-the counter or from other sources
Thyroid hormone status defines brown adipose tissue activity and browning of white adipose tissues in mice
The present study aimed to determine the effect of thyroid hormone dysfunction
on brown adipose tissue activity and white adipose tissue browning in mice.
Twenty randomized female C57BL/6NTac mice per treatment group housed at room
temperature were rendered hypothyroid or hyperthyroid. In-vivo small animal
18F-FDG PET/MRI was performed to determine the effects of hypo- and
hyperthyroidism on BAT mass and BAT activity. Ex-vivo14C-acetate loading assay
and assessment of thermogenic gene and protein expression permitted analysis
of oxidative and thermogenic capacities of WAT and BAT of eu-, hyper and
hypothyroid mice. 18F-FDG PET/MRI revealed a lack of brown adipose tissue
activity in hypothyroid mice, whereas hyperthyroid mice displayed increased
BAT mass alongside enhanced 18F-FDG uptake. In white adipose tissue of both,
hyper- and hypothyroid mice, we found a significant induction of thermogenic
genes together with multilocular adipocytes expressing UCP1. Taken together,
these results suggest that both the hyperthyroid and hypothyroid state
stimulate WAT thermogenesis most likely as a consequence of enhanced
adrenergic signaling or compensation for impaired BAT function, respectively
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3,5-T2-A Janus-Faced Thyroid Hormone Metabolite Exerts Both Canonical T3-Mimetic Endocrine and Intracrine Hepatic Action.
Over the last decades, thyroid hormone metabolites (THMs) received marked attention as it has been demonstrated that they are bioactive compounds. Their concentrations were determined by immunoassay or mass-spectrometry methods. Among those metabolites, 3,5-diiodothyronine (3,5-T2), occurs at low nanomolar concentrations in human serum, but might reach tissue concentrations similar to those of T4 and T3, at least based on data from rodent models. However, the immunoassay-based measurements in human sera revealed remarkable variations depending on antibodies used in the assays and thus need to be interpreted with caution. In clinical experimental approaches in euthyroid volunteers and hypothyroid patients using the immunoassay as the analytical tool no evidence of formation of 3,5-T2 from its putative precursors T4 or T3 was found, nor was any support found for the assumption that 3,5-T2 might represent a direct precursor for serum 3-T1-AM generated by combined deiodination and decarboxylation from 3,5-T2, as previously documented for mouse intestinal mucosa. We hypothesized that lowered endogenous production of 3,5-T2 in patients requiring T4 replacement therapy after thyroidectomy or for treatment of autoimmune thyroid disease, compared to production of 3,5-T2 in individuals with intact thyroid glands might contribute to the discontent seen in a subset of patients with this therapeutic regimen. So far, our observations do not support this assumption. However, the unexpected association between high serum 3,5-T2 and elevated urinary concentrations of metabolites related to coffee consumption requires further studies for an explanation. Elevated 3,5-T2 serum concentrations were found in several situations including impaired renal function, chronic dialysis, sepsis, non-survival in the ICU as well as post-operative atrial fibrillation (POAF) in studies using a monoclonal antibody-based chemoluminescence immunoassay. Pilot analysis of human sera using LC-linear-ion-trap-mass-spectrometry yielded 3,5-T2 concentrations below the limit of quantification in the majority of cases, thus the divergent results of both methods need to be reconciliated by further studies. Although positive anti-steatotic effects have been observed in rodent models, use of 3,5-T2 as a muscle anabolic, slimming or fitness drug, easily obtained without medical prescription, must be advised against, considering its potency in suppressing the HPT axis and causing adverse cardiac side effects. 3,5-T2 escapes regular detection by commercially available clinical routine assays used for thyroid function tests, which may be seriously disrupted in individuals self-administering 3,5-T2 obtained over-the counter or from other sources
3,5-T2—A Janus-Faced Thyroid Hormone Metabolite Exerts Both Canonical T3-Mimetic Endocrine and Intracrine Hepatic Action
Over the last decades, thyroid hormone metabolites (THMs) received marked attention as it has been demonstrated that they are bioactive compounds. Their concentrations were determined by immunoassay or mass-spectrometry methods. Among those metabolites, 3,5-diiodothyronine (3,5-T2), occurs at low nanomolar concentrations in human serum, but might reach tissue concentrations similar to those of T4 and T3, at least based on data from rodent models. However, the immunoassay-based measurements in human sera revealed remarkable variations depending on antibodies used in the assays and thus need to be interpreted with caution. In clinical experimental approaches in euthyroid volunteers and hypothyroid patients using the immunoassay as the analytical tool no evidence of formation of 3,5-T2 from its putative precursors T4 or T3 was found, nor was any support found for the assumption that 3,5-T2 might represent a direct precursor for serum 3-T1-AM generated by combined deiodination and decarboxylation from 3,5-T2, as previously documented for mouse intestinal mucosa. We hypothesized that lowered endogenous production of 3,5-T2 in patients requiring T4 replacement therapy after thyroidectomy or for treatment of autoimmune thyroid disease, compared to production of 3,5-T2 in individuals with intact thyroid glands might contribute to the discontent seen in a subset of patients with this therapeutic regimen. So far, our observations do not support this assumption. However, the unexpected association between high serum 3,5-T2 and elevated urinary concentrations of metabolites related to coffee consumption requires further studies for an explanation. Elevated 3,5-T2 serum concentrations were found in several situations including impaired renal function, chronic dialysis, sepsis, non-survival in the ICU as well as post-operative atrial fibrillation (POAF) in studies using a monoclonal antibody-based chemoluminescence immunoassay. Pilot analysis of human sera using LC-linear-ion-trap-mass-spectrometry yielded 3,5-T2 concentrations below the limit of quantification in the majority of cases, thus the divergent results of both methods need to be reconciliated by further studies. Although positive anti-steatotic effects have been observed in rodent models, use of 3,5-T2 as a muscle anabolic, slimming or fitness drug, easily obtained without medical prescription, must be advised against, considering its potency in suppressing the HPT axis and causing adverse cardiac side effects. 3,5-T2 escapes regular detection by commercially available clinical routine assays used for thyroid function tests, which may be seriously disrupted in individuals self-administering 3,5-T2 obtained over-the counter or from other sources
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