A pediatric hypophosphatemic rickets on MRI, 99mTc-MDP bone scan and 18F-FDG PET/CT.

We present a case of a 13 years old boy who was hospitalized with a 10 months history of progressive pain and weakness in his lower extremities. The laboratory tests revealed slightly decreased phosphate and 25-hydroxyvitamin D3, high alkaline phosphatase, normal calcium and parathyroid hormone (PTH). Magnetic resonance imaging (MRI) showed multiple patchy lesions indicating bone destruction in the metaphyses and epiphyses of the left knee. Fluorine-18-fluorodeoxyglucose positron emission tomography/computed tomography (18F-FDG PET/CT) revealed a generalized decrease of bone density in axial bones with slightly increased 18F-FDG metabolism. Whole body technetium-99m methylene diphosphonate (99mTc-MDP) scintigraphy revealed multiple areas of increased uptake at costochondral junctions of the ribs bilaterally suggesting a rachitic rosary and at the metaphyses of the bones of the limbs. Based on these findings we suggested the diagnosis of hypophosphatemic rickets (HPR). Phosphate and vitamin D substitution resulted in clinical improvement of the symptoms after 3 months.

Effect of hypoparathyroid on bone mineral density of lumber spine in postmenopausal women with differentiated thyroid cancinoma.

OBJECTIVE: To investigate the effect of hypoparathyroid on bone mineral density in postmenopausal women with differentiated thyroid carcinoma (DTC). SUBJECT AND METHODS: Postmenopausal women with postoperative DTC, and undergoing thyroid residual ablation or for metastases treatment were collected and followed for two years. They were divided into hypoparathyroid group (PTH<15pg/mL) and a normal cognitive group (PTH>15pg/mL). Bone mineral density (BMD) at the lumbar spine was analyzed using dual-energy X-ray absorptiometry (DXA) at baseline, 6, 12 and 24 months. All patients had calcium and active vitamin D supplementation. RESULTS: The thyroid cancers included 211 papillary carcinomas, 14 follicular carcinomas. The majority of them were retired from previous work (157/225, 70%). There were 45 DTC patients in hypoparathyroid group and 180 patients in PTH normal group (postmenopausal controls). They are comparable in age, TSH suppression, BMD at baseline. There is no significant difference in BMD of lumbar spine between hypoparathyroid group and postmenopausal controls at baseline 6, 12 and 24 months follow-up which were1.03±0.14 and 1.04±0.18 (t=0.4, P=0.69), 1.04±0.13 and 1.01±0.19 (t=1.25, P=0.21), 1.06±0.15 and 1.02±0.16 (t=1.16, P=0.26), 1.06±0.21 and 1.01±0.17 (t=0.93, P=0.29), respectively. Areal BMD was increased by 2.9% in hypoparathyroid group in the lumbar spine at 12 and 24 months follow-up, while decrease of 2.9% in postmenopausal controls. No increase in BMD at lumbar spine was found in postmenopausal controls. CONCLUSION: Transient hypoparathyroid increased BMD at lumbar spine by DXA in postmenopausal DTC patients compared to postmenopausal controls.

A pediatric malignant paraganglioma and brief review of the literature.

A 10 years old boy presented to our hospital complaining of belly pain. He had a retroperitoneal mass diagnosed by ultrasound 3 days before. During that time he was found to have slight hypertension. Computed tomography (CT) showed a left retroperitoneal mass with edge enhancement and central necrosis indicating pheochromocytoma (PCC). Vanillymandelic acid (VMA) in the 24 hours urine sample was not elevated. Pheochromocytoma was suspected given his hypertension and the ultrasound and CT findings. Fluorine-18- fluorodeoxy glucose positron emission tomography (18F-FDG PET) showed intense uptake in the left adrenal area (SUVmax 32.9) with a central 18F-FDG uptake defect. Subsequently, left adrenalectomy was successfully performed. Histological examination showed that the tumor was a paraganglioma (PGL) with low-grade malignancy. CONCLUSION: Fluorine-18-FDG PET is a highly sensitive method to detect PGL, but could not make a differential diagnosis between PGL and PCC although high uptake of 18F-FDG may indicate malignancy. As our case clearly demonstrates, rare cases of PCC or PGL should be considered in the setting of secondary hypertension.

Massively parallel sequencing identifies the gene Megf8 with ENU-induced mutation causing heterotaxy.

Forward genetic screens with ENU (N-ethyl-N-nitrosourea) mutagenesis can facilitate gene discovery, but mutation identification is often difficult. We present the first study in which an ENU-induced mutation was identified by massively parallel DNA sequencing. This mutation causes heterotaxy and complex congenital heart defects and was mapped to a 2.2-Mb interval on mouse chromosome 7. Massively parallel sequencing of the entire 2.2-Mb interval identified 2 single-base substitutions, one in an intergenic region and a second causing replacement of a highly conserved cysteine with arginine (C193R) in the gene Megf8. Megf8 is evolutionarily conserved from human to fruit fly, and is observed to be ubiquitously expressed. Morpholino knockdown of Megf8 in zebrafish embryos resulted in a high incidence of heterotaxy, indicating a conserved role in laterality specification. Megf8(C193R) mouse mutants show normal breaking of symmetry at the node, but Nodal signaling failed to be propagated to the left lateral plate mesoderm. Videomicroscopy showed nodal cilia motility, which is required for left-right patterning, is unaffected. Although this protein is predicted to have receptor function based on its amino acid sequence, surprisingly confocal imaging showed it is translocated into the nucleus, where it is colocalized with Gfi1b and Baf60C, two proteins involved in chromatin remodeling. Overall, through the recovery of an ENU-induced mutation, we uncovered Megf8 as an essential regulator of left-right patterning.

ApoA-1 mimetic peptide reverses uremia-induced upregulation of pro-atherogenic pathways in the aorta.

BACKGROUND: Chronic kidney disease (CKD) results in accelerated atherosclerosis and cardiovascular disease. This is primarily mediated by oxidative stress, inflammation and dyslipidemia. By mediating reverse cholesterol transport and exerting antioxidant/anti-inflammatory actions, high-density lipoprotein (HDL) and ApoA-1 protect against atherosclerosis. Plasma Apo-1, HDL cholesterol and HDL antioxidant/anti-inflammatory activities are reduced in CKD. ApoA-1 mimetic peptides associate with and enhance antioxidant/anti-inflammatory properties of HDL. We hypothesized that long-term administration of ApoA-1 mimetic peptide, L4F, may ameliorate inflammation and oxidative stress in the conduit arteries in experimental CKD. METHODS: After 5/6 nephrectomy, rats were randomized to L4F (5 mg/kg s.c. 3 times weekly for 4 weeks) and placebo-treated groups. Sham-operated rats served as controls. RESULTS: The untreated CKD group exhibited marked lipid accumulation and upregulations of NAD(P)H oxidase subunits (gp91(phox), p22(phox), and p47(phox)), COX-2, 12-lipoxygenase, MCP-1, PAI-1, myeloperoxidase and iNOS, NFκB activation and nitrotyrosine accumulation in the thoracic aorta. L4F administration reversed or attenuated these abnormalities without altering renal function or plasma lipids. CONCLUSIONS: CKD leads to lipid accumulation and upregulation of pro-atherogenic pathways in the artery wall. These abnormalities are attenuated by ApoA-1 mimetic peptide, pointing to its protective effect in CKD. Future studies are needed to explore the effect of these peptides in CKD patients.

FDG PET Predicts the Effects of 131I and Prognosis for Patients with Bone Metastases from Differentiated Thyroid Carcinoma.

BACKGROUND: 18F-FDG PET and 131I scans are important in the detection of metastases from differentiated thyroid carcinoma (DTC). The relationship of FDG and radioiodine (RAI) metabolism in bone metastases (BMs) from DTC and its prognostic value on RAI treatment is not clear. METHODS: The retrospective study included DTC patients with BMs from two medical centers, who underwent 18F-FDG PET/CT scans and RAI therapy. Therapeutic response was evaluated by serum thyroglobulin (Tg) levels and anatomical imaging changes. RESULTS: The analyses were performed on 30 patients with 72 BMs. Forty-two (42/72, 58%) lesions displayed simultaneous 131I and 18F-FDG uptake. BMs with positive 18F-FDG uptake had a less favorable response to RAI therapy in comparison to those with negative 18F-FDG uptake (p = 0.018), even in 131I-avid lesions (p = 0.033). Sixteen (53%) patients had disease progression with a median PFI of 26 months (range: 3 to 89 months). Compared to those with 131I-avid but non-FDG-avid BMs, patients presenting with 18F-FDG-avid BMs had shorter PFI, whether with 131I uptake (p = 0.002) or without (p = 0.002). CONCLUSION: More than half of BMs (58%) from DTC show simultaneous 18F-FDG and 131I uptakes which are contrary to the flip-flop phenomenon (131I negative and 18F-FDG positive). Regardless of 131I uptake, 18F-FDG uptake of BMs portends a less favorable therapeutic response and poorer prognosis for patients with DTC.

Regional expression of NAD(P)H oxidase and superoxide dismutase in the brain of rats with neurogenic hypertension.

BACKGROUND: Single injection of small quantities of phenol into the kidney cortex causes hypertension which is mediated by renal afferent sympathetic pathway activation. This phenomenon can be prevented by superoxide dismutase (SOD) infusion in the lateral ventricle, suggesting the role of superoxide (O(2)(-).) in noradrenergic control of arterial pressure. Since NAD(P)H oxidase is a major source of O(2)(-)., we tested the hypothesis that hypertension in this model may be associated with upregulation of NAD(P)H oxidase in relevant regions of brain. METHODS: NAD(P)H oxidase subunits, mitochondrial (MnSOD) and cytoplasmic (CuZnSOD) SOD were measured in rats 4 weeks after injection of phenol or saline in the left kidney cortex. RESULTS: Phenol-injected rats exhibited hypertension, upregulation of gp91(phox), p22(phox), p47(phox) and p67(phox) in the medulla, gp91(phox) and p22(phox) in pons and gp91(phox) in hypothalamus. This was associated with upregulation of MnSOD with little change in CuZnSOD. CONCLUSIONS: Chronic hypertension in phenol-injected rats is associated with upregulation of NAD(P)H oxidase and hence increased O(2)(-). production capacity in the key regions of the brain involved in regulation of blood pressure. Since reactive oxygen species can intensify central noradrenergic activity, the observed maladaptive changes may contribute to the genesis and maintenance of the associated hypertension.

Regional expression of NO synthase, NAD(P)H oxidase and superoxide dismutase in the rat brain.

Nitric oxide (NO) derived from the endothelial NO synthase (eNOS) contributes to regulation of cerebral circulation, whereas that produced by neuronal NOS (nNOS) participates in the regulation of brain function. In particular, NO plays an important role in modulation of sympathetic activity and hence central regulation of arterial pressure. Superoxide derived from NAD(P)H oxidase avidly reacts with and inactivates NO and, thereby, modulates its bioavailability. Calmodulin (CM) is required for activation of NOS and soluble guanylate cyclase (sGC) serves as a NO receptor. Superoxide is dismutated to H2O2 by superoxide dismutase (SOD) and H2O2 is converted to H2O by catalase or glutathione peroxidase (GPX). Given the importance of NO in the regulation of brain perfusion and function, we undertook the present study to determine the relative expressions of immunodetectable nNOS, eNOS, CM, sGC, NAD(P)H oxidase and SOD by Western analysis in different regions of the normal rat brain. nNOS was abundantly expressed in the pons cerebellum and hypothalamus and less so in the cortex and medulla. sGC abundance was highest in the hypothalamus and pons, and lowest in the cerebellum and medulla. eNOS and calmodulin were equally abundant in all regions. NAD(P)H oxide was most abundant in the pons compared to other regions. Cytoplasmic SOD was equally distributed among different regions but catalase and GPX were more abundant in pons, hypothalamus and medulla and less so in the cortex and cerebellum. Thus, the study documented regional distributions of NOS, NAD(P)H oxidase, antioxidant enzymes, sGC and calmodulin which collectively regulate production and biological activities of NO and superoxide, the two important small molecular size signaling molecules.

Telomere instability in a human tumor cell line expressing NBS1 with mutations at sites phosphorylated by ATM.

Nijmegen breakage syndrome (NBS) is an autosomal genetic disease demonstrating a variety of phenotypic abnormalities, including premature aging, increased cancer incidence, chromosome instability, and sensitivity to ionizing radiation. The gene involved in NBS, NBS1, is part of the MRE11/RAD50/NBS1 (MRN) complex that also includes MRE11 and RAD50, which is involved in DNA repair and cell cycle regulation in response to DNA damage. The MRN complex is also involved in telomere maintenance, as demonstrated by the shortened telomeres in NBS primary human fibroblasts and the association of NBS1 with the telomere-binding protein TRF2. To learn more about how a deficiency in telomere maintenance might contribute to chromosome instability in NBS, we have investigated the stability of telomeres in two telomerase-positive human tumor cell clones, BNmt-On and BNmt-Off, expressing an inducible NBS1(S278A/S343A) gene containing mutations at serines 278 and 343 phosphorylated by ATM. The results demonstrate an increased rate of telomere loss in both clones following expression of NBS1(S278A/S343A). The absence of detectable changes in average telomere length suggests that NBS1-associated telomere loss results from stochastic events involving complete telomere loss or loss of telomere capping function. The recombination events associated with telomere loss were found to be similar to those shown previously to result in breakage/fusion/bridge cycles, suggesting that telomere loss can contribute to chromosome instability in NBS1-deficient cells. Telomere loss showed no correlation with radiosensitivity or radioresistant DNA synthesis, demonstrating that NBS1(S278A/S343A) promotes telomere loss through a separate pathway from these other phenotypes associated with NBS.

Isoform-selective interactions between estrogen receptors and steroid receptor coactivators promoted by estradiol and ErbB-2 signaling in living cells.

Estrogen receptor (ER)alpha and -beta interact with a variety of coactivator proteins, most notably members of the steroid receptor coactivator (SRC) family, and these interactions have been shown to be regulated by estrogenic ligands and growth factor signaling. Here, using fluorescence resonance energy transfer (FRET), the selectivity of different stimulants on ERalpha and -beta interactions with coactivator receptor interaction domains (RIDs) were examined in living cells. We first show that ERalpha and ERbeta homo- and heterodimers form in vivo independently of the presence of 17beta-estradiol (E2) or antiestrogens. We then demonstrate that E2 enhances interactions between ERalpha and the RIDs of SRC-1 and SRC-3, whereas the interaction between ERalpha with the SRC-2 RID is ligand independent. The transcriptionally inactive mutant ERalphaL539A showed no interaction with all three SRC RIDs. Similarly, treatment with the antagonists 4-hydroxytamoxifen and EM-652 abolished all interactions between ERalpha and the SRC RIDs. FRET data also demonstrate that, in contrast to ERalpha, ERbeta interacts with all three SRC RIDs in a ligand-independent manner. However, these interactions were further enhanced or stabilized by E2, whereas the antiestrogen EM-652 abolished all interactions. In the presence of both ERalpha and ERbeta, E2 treatment led to the recruitment of SRC RIDs to the nuclei. Finally, expression of the oncogenic activated ErbB-2/Neu protein specifically enhanced ERalpha but not ERbeta interactions with SRC RIDs to an extent similar to E2-stimulated interactions. In summary, using FRET, we demonstrated preferential interactions between ER isoforms and coactivators upon hormonal treatment and activation of a growth factor signal transduction pathway in living cells.

Telomere instability in a human tumor cell line expressing a dominant-negative WRN protein.

Werner Syndrome (WS) is an autosomal recessive disease characterized by premature aging and chromosome instability. The protein involved in WS, WRN, is a RecQ-type helicase that also has exonuclease activity. WRN has been demonstrated to bind to a variety of other proteins, including RPA, DNA-PKcs, and TRF2, suggesting that WRN is involved in DNA replication, repair, recombination, and telomere maintenance. In culture, WS cells show premature senescence, which can be overcome by transfection with an expression vector containing the gene for the catalytic subunit of telomerase. However, telomerase expression does not eliminate chromosome instability in WS cells, which led to the proposal that telomere loss is not the cause of the high rate of chromosome rearrangements in WS cells. In the present study, we have investigated how a WRN protein containing a dominant-negative mutation (K577M-WRN) influences the stability of telomeres in a human tumor cell line expressing telomerase. The results demonstrate an increased rate of telomere loss and chromosome fusion in cells expressing K577M-WRN. Expression of K577M-WRN results in reduced levels of telomerase activity, however, the absence of detectable changes in average telomere length demonstrates that WRN-associated telomere loss results from stochastic events involving complete telomere loss or loss of telomere capping function. Thus, telomere loss can contribute to chromosome instability in cells deficient in WRN regardless of the expression of telomerase activity.

Itch E3 ligase-mediated regulation of TGF-beta signaling by modulating smad2 phosphorylation.

Protein ubiquitination has been implicated in the intracellular biochemical events transduced by TGF-beta receptor via different mechanisms including the degradation of Smads or their binding proteins. Here we show that loss of Itch E3 ligase in mouse embryonic fibroblasts (MEFs) results in reduced susceptibility of TGF-beta-induced cell growth arrest and decreased phosphorylation of Smad2, without apparent alteration in protein levels for Smad2, Smad4, and Smad7 in Itch-/- MEFs. Itch promotes ubiquitination of Smad2 and augments Smad2 phosphorylation that requires an intact ligase activity of Itch. Moreover, Itch facilitates complex formation between TGF-beta receptor and Smad2 and enhances TGF-beta-induced transcription. This study reveals a previously unrecognized positive TGF-beta signaling pathway via proteolysis-independent ubiquitination.