Enhanced re-myelination in transthyretin null mice following cuprizone mediated demyelination.

Thyroid hormones (THs) impact nearly every tissue in the body, including the adult and developing central nervous system. The distribution of THs around the body is facilitated by specific TH distributor proteins including transthyretin (TTR). In addition to being produced in the liver, TTR is synthesized in the choroid plexus of the brain. The synthesis of TTR by choroid plexus epithelial cells allows transport of THs from the blood into the brain. Adequate supply of THs to the brain is required for developmental myelination of axons and the maintenance of mature myelin throughout adult life, essential for the proper conduction of nerve impulses. Insufficient THs in developing mice results in hypo-myelination (thinner myelin around axons). However, confounding evidence demonstrated that in developing brain of TTR null mice, hyper-myelination of axons was observed in the corpus callosum. This raised the question whether increased myelination occurs during re-myelination in the adult brain following targeted demyelination. To investigate the effect of TTR during re-myelination, cuprizone induced depletion of myelin in the corpus callosum of adult mice was initiated, followed by a period of myelin repair. Myelin thickness was measured to assess re-myelination rates for 6 weeks. TTR null mice displayed expedited rates of early re-myelination, preferentially re-myelinating smaller axons compared to those of wild type mice. Furthermore, TTR null mice produced thicker myelin than wild type mice during re-myelination. These results may have broader implications in understanding mechanisms governing re-myelination, particularly in potential therapeutic contexts for acquired demyelinating diseases such as multiple sclerosis.

Gender-specific effects of transthyretin on neural stem cell fate in the subventricular zone of the adult mouse.

Choroid plexus epithelial cells produce and secrete transthyretin (TTR). TTR binds and distributes thyroid hormone (TH) to brain cells via the cerebrospinal fluid. The adult murine subventricular zone (SVZ) is in close proximity to the choroid plexus. In the SVZ, TH determines neural stem cell (NSC) fate towards a neuronal or a glial cell. We investigated whether the loss of TTR also disrupted NSC fate choice. Our results show a decreased neurogenic versus oligodendrogenic balance in the lateroventral SVZ of Ttr knockout mice. This balance was also decreased in the dorsal SVZ, but only in Ttr knockout male mice, concomitant with an increased oligodendrocyte precursor density in the corpus callosum. Quantitative RTqPCR analysis following FACS-dissected SVZs, or marked-coupled microbeads sorting of in vitro neurospheres, showed elevated Ttr mRNA levels in neuronal cells, as compared to uncommitted precursor and glial cells. However, TTR protein was undetectable in vivo using immunostaining, and this despite the presence of Ttr mRNA-expressing SVZ cells. Altogether, our data demonstrate that TTR is an important factor in SVZ neuro- and oligodendrogenesis. They also reveal important gender-specific differences and spatial heterogeneity, providing new avenues for stimulating endogenous repair in neurodegenerative diseases.

Assessment of patients with hereditary transthyretin amyloidosis - understanding the impact of management and disease progression.

Timely diagnosis of hereditary variant transthyretin (ATTRv) amyloidosis is critical for appropriate treatment and optimal outcomes. Significant differences are seen between patients receiving treatment and those who are not, though disease progression may continue despite treatment in some patients. Healthcare professionals caring for patients with ATTRv amyloidosis therefore need reliable ongoing assessments to understand the continuing course of disease and make appropriate treatment choices on an individual basis. Various signs and symptoms experienced by patients may be evaluated as indicators of disease progression, though there is currently no validated score that can be used for such ongoing assessment. Recognizing this situation, a group of clinicians highly experienced in ATTR amyloidosis developed an approach to understand and define disease progression in diagnosed and treated patients with ATTRv amyloidosis. The suggested approach is based on the recognition of distinct phenotypes which may usefully inform the particular tools, tests and investigations that are most likely to be appropriate for individual patients. It is aimed at implementing appropriate and ongoing assessment of patients being treated for ATTRv amyloidosis, such that the effectiveness of management can be usefully assessed throughout the course of disease and management can be tailored according to the patient's requirements.

Hepatocyte-specific delivery of siRNAs conjugated to novel non-nucleosidic trivalent N-acetylgalactosamine elicits robust gene silencing in vivo.

We recently demonstrated that siRNAs conjugated to triantennary N-acetylgalactosamine (GalNAc) induce robust RNAi-mediated gene silencing in the liver, owing to uptake mediated by the asialoglycoprotein receptor (ASGPR). Novel monovalent GalNAc units, based on a non-nucleosidic linker, were developed to yield simplified trivalent GalNAc-conjugated oligonucleotides under solid-phase synthesis conditions. Synthesis of oligonucleotide conjugates using monovalent GalNAc building blocks required fewer synthetic steps compared to the previously optimized triantennary GalNAc construct. The redesigned trivalent GalNAc ligand maintained optimal valency, spatial orientation, and distance between the sugar moieties for proper recognition by ASGPR. siRNA conjugates were synthesized by sequential covalent attachment of the trivalent GalNAc to the 3'-end of the sense strand and resulted in a conjugate with in vitro and in vivo potency similar to that of the parent trivalent GalNAc conjugate design.

Silencing of murine transthyretin and retinol binding protein genes has distinct and shared behavioral and neuropathologic effects.

The murine genes encoding transthyretin (TTR) and retinol binding protein (RBP) were independently silenced by targeted disruption more than 10 years ago. Studies of both strains showed surprisingly little impact on either thyroid function or retinoid metabolism. Silencing TTR led to a relatively mild behavioral phenotype. In order to gain insight into the behavioral effect and determine if it was related to TTR's function as the carrier of RBP we carried out simultaneous studies with homozygous Rbp4(-/-) and Ttr(-/-) animals 4-7 months of age. Both strains showed behavioral differences relative to Ttr and Rbp4 wild-type animals and each other. The patterns were discrete for each knockout although there was some overlap. Neuropathologic examination of the cortex and hippocampus revealed cortical and hippocampal (CA3) neuronal loss in both and some degree of gliosis, more pronounced in the Rbp4(-/-) mice. There also appeared to be a major reduction in proliferating neuroblasts in the subventricular zone in both strains, which was also more severe in the Rbp4(-/-) mice. This is the first description of behavioral abnormalities in Rbp4(-/-)mice. The data also indicate that it is unlikely that the behaviors seen in Ttr(-/-) mice are related to its function as an RBP carrier.

Neuronal production of transthyretin in human and murine Alzheimer's disease: is it protective?

Transthyretin (TTR), a systemic amyloid precursor in the human TTR amyloidoses, interacts with ß-amyloid (Aß) in vitro, inhibits Aß fibril formation, and suppresses the Alzheimer's disease (AD) phenotype in APP23 mice bearing a human APP gene containing the Swedish autosomal dominant AD mutation. In the present study, we show that TTR is a neuronal product upregulated in AD. Immunohistochemical analysis reveals that, in contrast to brains from non-demented age-matched individuals and control mice, the majority of hippocampal neurons from human AD and all those from the APP23 mouse brains contain TTR. Quantitative PCR for TTR mRNA and Western blot analysis show that primary neurons from APP23 mice transcribe TTR mRNA, and the cells synthesize and secrete TTR protein. TTR mRNA abundance is greatly increased in cultured cortical and hippocampal embryonic neurons and cortical lysates from adult APP23 mice. Antibodies specific for TTR and Aß pulled down TTR/Aß complexes from cerebral cortical extracts of APP23 mice and some human AD patients but not from control brains. In complementary tissue culture experiments, recombinant human TTR suppressed the cytotoxicity of soluble Aß aggregates added to mouse neurons and differentiated human SH-SY5Y neuroblastoma cells. The findings that production of Aß, its precursor, or its related peptides induces neuronal TTR transcription and synthesis and the presence of Aß/TTR complexes in vivo suggest that increased TTR production coupled with interaction between TTR and Aß and/or its related peptides may play a role in natural resistance to human AD.

Gender-dependent transthyretin modulation of brain amyloid-ß levels: evidence from a mouse model of Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disorder affecting tens of millions of people worldwide, with women being at greater risk of developing the disease. A growing body of evidence suggests transthyretin (TTR) as an important modulator of AD pathogenesis. Aiming at providing further insight into the potential neuroprotective role of TTR and gender differences in AD, we crossed transgenic AßPPswe/PS1A246E mice with TTR-null mice and investigated both male and female AßPPswe/PS1A246E/TTR+/+, AßPPswe/PS1A246E/TTR+/-, and AßPPswe/PS1A246E/TTR-/- animals for brain amyloid-ß (Aß) levels and deposition. The levels of circulating TTR between non-transgenic and AD mice were evaluated. Decreased levels of circulating TTR in AD mice as compared to non-transgenic littermates were observed in early stages of AD-like neuropathology, but not at later stages where an opposite relationship was found. Elevated brain levels of Aß42 were observed in AßPPswe/PS1A246E/TTR+/- female mice as compared to AßPPswe/PS1A246E/TTR+/+ female littermates; no significant differences were found among males of different TTR genotypes. We subsequently quantified the brain levels of testosterone and 17ß-estradiol in these animals and verified that AßPPswe/PS1A246E/TTR+/- female mice present reduced brain levels of both hormones as compared to AßPPswe/PS1A246E/TTR+/+ females; no significant differences were detected among males of different TTR genotypes. Our results provide evidence for a gender-associated modulation of brain Aß levels and brain sex steroid hormones by TTR, and suggest that reduced levels of brain testosterone and 17ß-estradiol in female mice with TTR genetic reduction might underlie their increased AD-like neuropathology.

Possible involvement of transthyretin in hippocampal beta-amyloid burden and learning behaviors in a mouse model of Alzheimer's disease (TgCRND8).

BACKGROUND: Alzheimer's disease (AD) is a neurodegenerative disease characterized by progressive memory loss, possibly triggered by the accumulation of beta-amyloid (Abeta) peptides and the hyperphosphorylation of Tau neurofilament protein. Recent findings have shown that transthyretin (TTR) is a potent scavenger of Abeta peptide deposits, suggesting a possible neuroprotective role for TTR in neurodegenerative processes associated with amyloidogenesis, such as AD. METHODS: To investigate the relationship between TTR and Abeta deposition, we crossed mouse carrying a deletion of TTR (TTR(- or -)) with a transgenic mouse model of AD (TgCRND8), and Abeta burden and spatial learning capacities were evaluated at 4 and 6 months of age (exclusion of the 6 month-old TgCRND8/TTR(- or -) group due to low survival rate). RESULTS: Rather surprisingly, Abeta plaque burden was significantly reduced in the hippocampus of 4-month-old TgCRND8/TTR(+ or -), and to a lesser extent in TgCRND8/TTR(- or -), as compared to age-matched TgCRND8/TTR(+ or +). No difference in plaque burden was found between any groups in 6-month-old animals. At 4 and 6 months of age, all populations of these hybrid transgenic mice displayed similar magnitude of spatial memory deficits in the Morris water maze task. CONCLUSION: Since TgCRND8 mice represent an aggressive model of Abeta deposition with plaques developing as early as 3 months of age, along with spatial learning deficits, it may be already too late at 4 and 6 months of age to observe significant changes due to the deletion of the TTR gene.

Neuropeptide Y expression and function during osteoblast differentiation--insights from transthyretin knockout mice.

To better understand the role of neuropeptide Y (NPY) in bone homeostasis, as its function in the regulation of bone mass is unclear, we assessed its expression in this tissue. By immunohistochemistry, we demonstrated, both at embryonic stages and in the adult, that NPY is synthesized by osteoblasts, osteocytes, and chondrocytes. Moreover, peptidylglycine alpha-amidating monooxygenase, the enzyme responsible for NPY activation by amidation, was also expressed in these cell types. Using transthyretin (TTR) KO mice as a model of augmented NPY levels, we showed that this strain has increased NPY content in the bone, further validating the expression of this neuropeptide by bone cells. Moreover, the higher amidated neuropeptide levels in TTR KO mice were related to increased bone mineral density and trabecular volume. Additionally, RT-PCR analysis established that NPY is not only expressed in MC3T3-E1 osteoblastic cells and bone marrow stromal cells (BMSCs), but is also detectable by RIA in BMSCs undergoing osteoblastic differentiation. In agreement with our in vivo observations, in vitro, TTR KO BMSCs differentiated in osteoblasts had increased NPY levels and exhibited enhanced competence in undergoing osteoblastic differentiation. In summary, this work contributes to a better understanding of the role of NPY in the regulation of bone formation by showing that this neuropeptide is expressed in bone cells and that increased amidated neuropeptide content is related to increased bone mass.

Chapter 17: Transthyretin: an enhancer of nerve regeneration.

Transthyretin (TTR), a plasma and cerebrospinal fluid protein secreted by the liver and choroid plexus, is mainly known as the physiological carrier of thyroxine (T(4)) and retinol. Under pathological conditions, various TTR mutations are related to familial amyloid polyneuropathy (FAP), a neurodegenerative disorder characterized by deposition of TTR amyloid fibrils, particularly in the peripheral nervous system (PNS), leading to axonal loss and neuronal death. Recently, a number of TTR functions in neurobiology have been described; these may explain the preferential TTR deposition, when mutated, in the PNS of FAP patients. In this respect, and with a particular relevance in the PNS, TTR has been shown to have the ability to enhance neurite outgrowth in vitro and nerve regeneration following injury, in vivo. In the following pages, this novel TTR function, as well as its importance in nerve biology and repair will be discussed.

Transthyretin internalization by sensory neurons is megalin mediated and necessary for its neuritogenic activity.

Mutated transthyretin (TTR) causes familial amyloid polyneuropathy, a neurodegenerative disorder characterized by TTR deposition in the peripheral nervous system (PNS). The origin/reason for TTR deposition in the nerve is unknown. Here we demonstrate that both endogenous mouse TTR and TTR injected intravenously have access to the mouse sciatic nerve. We previously determined that in the absence of TTR, both neurite outgrowth in vitro and nerve regeneration in vivo were impaired. Reinforcing this finding, we now show that local TTR delivery to the crushed sciatic nerve rescues the regeneration phenotype of TTR knock-out (KO) mice. As the absence of TTR was unrelated to neuronal survival, we further evaluated the Schwann cell and inflammatory response to injury, as well as axonal retrograde transport, in the presence/absence of TTR. Only retrograde transport was impaired in TTR KO mice which, in addition to the neurite outgrowth impairment, might account for the decreased regeneration in this strain. Moreover, we show that in vitro, in dorsal root ganglia neurons, clathrin-dependent megalin-mediated TTR internalization is needed for TTR neuritogenic activity. Supporting this observation, we demonstrate that in vivo, decreased levels of megalin lead to decreased nerve regeneration and that megalin's action as a regeneration enhancer is dependent on TTR. In conclusion, our work unravels the mechanism of TTR action during nerve regeneration. Additionally, TTR presence in the nerve, as is here shown, may underlie its preferential deposition in the PNS of familial amyloid polyneuropathy patients.

Transthyretin accelerates vascular Abeta deposition in a mouse model of Alzheimer's disease.

Transthyretin (TTR) binds amyloid-beta (Abeta) and prevents Abeta fibril formation in vitro. It was reported that the lack of neurodegeneration in a transgenic mouse model of Alzheimer's disease (AD) (Tg2576 mouse) was associated with increased TTR level in the hippocampus, and that chronic infusion of anti-TTR antibody into the hippocampus of Tg2576 mice led to increased local Abeta deposits, tau hyperphosphorylation and apoptosis. TTR is, therefore, speculated to prevent Abeta pathology in AD. However, a role for TTR in Abeta deposition is not yet known. To investigate the relationship between TTR and Abeta deposition, we generated a mouse line carrying a null mutation at the endogenous TTR locus and the human mutant amyloid precursor protein cDNA responsible for familial AD (Tg2576/TTR(-/-) mouse) by crossing Tg2576 mice with TTR-deficient mice. We asked whether Abeta deposition was accelerated in Tg2576/TTR(-/-) mice relative to the heterozygous mutant Tg2576 (Tg2576/TTR(+/-)) mice. Contrary to our expectations, the degree of total and vascular Abeta burdens in the aged Tg2576/TTR(-/-) mice was significantly reduced relative to the age-matched Tg2576/TTR(+/-) mice. Our experiments present, for the first time, compelling evidence that TTR does not suppress but rather accelerates vascular Abeta deposition in the mouse model of AD.

The common environmental pollutant dioxin-induced memory deficits by altering estrogen pathways and a major route of retinol transport involving transthyretin.

Many toxic environmental and food agents have been suspected to be potential risk factors in inducing memory disabilities under normal and pathological conditions. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (known as dioxin or TCDD) is a common and prototypical member of a class of noxious environmental and food contaminants called the halogenated aromatic hydrocarbons. Since the role of dioxin in memory processes has not been studied in detail, the present report aims at elucidating the role of this pollutant in the maintenance of cognitive function. We found that TCDD (50miccrog/kg) induced spatial memory deficits in the Morris water maze (MWM) task in female but not male mice. This sex-dependant effect of dioxin seems to be related to the alteration of estrogen pathways, as treatment with 17beta-estradiol-3-benzoate (E; 5microg/day) reversed memory deficits induced by TCDD. We also observed that cognitive impairments produced by dioxin, which is known to interfere with retinoid turnover and metabolism, were abolished by retinoic acid (RA) treatment (150microg/kg). The cognitive effects of E and RA treatments seem to derive from common rather than additive mechanisms since memory deficits produced by TCDD were fully reversed by these compounds when used separately or in combination. Attenuation of dioxin-induced memory deficits in mice lacking transthyretin (TTR) suggests that TCDD may be acting by affecting the major route of retinol transport involving TTR. Taken together, these results suggest that the environmental and food pollutant TCDD can induce memory deficits by altering the estrogen pathways and a main route of TTR-mediated retinol transport.

[Nutritional status of adopted Chinese girls in Spain]. / El estado nutricional de niñas chinas adoptadas en España.

INTRODUCTION: Given the increasing number of adoptions from China in countries such as Spain, a study was designed to know and assess the nutritional profile at arrival and its likely implication in global health status, growth, and development of adopted Chinese girls. PATIENTS AND METHODS: Eighty-five Chinese girls adopted in Spain during the 2002-2003 period and ages ranging 7-33 months of life were studied at the Social Pediatrics Unit of the "Niño Jesús" Hospital, Madrid. Datas regarding institution of origin, nutritional parameters (anthropometrical and biochemical), presence of associated diseases, and developmental retardations (psychomotor/bone age) were gathered. RESULTS: Waterlow's nutritional index showed a malnourishment rate of 11% based on Chinese reference tables, as compared to 58% based on Spanish tables, of which 82% was acute malnourishment. When comparing the efficacy of both nutritional assessment methods proposed by Waterlow and Gomez, there were no differences in detection of malnourishment in spite of the fact that the latter author gives priority to the weight/age index as a nutritional indicator at ages lower than 2 years. Eighteen percent of the girls were considered at (height/age < p10) of suffering chronic malnourishment, but this was only confirmed in 67% of the group according to Waterlow's criteria. Among assessed plasma parameters, decreased prealbumin, lymphocytes, iron, and transferrin stand out. The predominant pathological findings were thalassemia, dermatitis, psychomotor retardation, and ferropenic anemia. CONCLUSIONS: These results make us believe in a possible improvement in the conditions at Chinese orphanages. The low age at the time of adoption may justify the low incidence of chronic nutritional deficiencies. As a whole, anthropometrical / nutritional impairments found may be related with the consequences of carelessness and vulnerability of the adoption process. In any case, early diagnosis and implementation of appropriate therapy as soon as possible, as it is being done, is essential to provide appropriate growth and development of these girls.

The absence of transthyretin does not impair regulation of lipid and glucose metabolism.

Increased levels of neuropeptide Y have been reported in transthyretin-null mice. This effect might be related to transthyretin ligands (retinol and thyroxine) since, through binding to nuclear receptors, they modulate the expression of genes that control cellular metabolism. The retinoic X receptors form obligatory heterodimers with peroxisome proliferator-activated receptors and liver X receptors - potent regulators of fat, glucose and cholesterol homeostasis. We used transthyretin-null mice to investigate whether the absence of transthyretin influences metabolism. Transthyretin-null mice do not differ from controls in body weight and white adipose tissue morphology, nor in basal or fast-induced circulating levels of glucose, lipids, and leptin. Glucose tolerance tests show that transthyretin-null mice have normal capacity to remove and metabolize energy substrates. Expression of genes encoding lipid transporters and nuclear receptors are also similar in transthyretin-null and control mice. Therefore, the absence of transthyretin does not seem to influence the regulation of lipid and glucose metabolism.

Cell division and apoptosis in the adult neural stem cell niche are differentially affected in transthyretin null mice.

Thyroid hormones (THs) are fundamental in regulation of growth and development, particularly of the brain. THs are required for full proliferative activity of neural stem cells in the subventricular zone (SVZ) of adult mouse brains, and also affect the normal fate of progenitor cells: apoptosis. Transthyretin (TTR) is a TH distributor protein in the blood and cerebrospinal fluid. TTR secretion by the choroid plexus is involved in transport of THs from blood into cerebrospinal fluid. We investigated the regulation of neural stem cell cycle in the SVZ of adult TTR null mice. Markers for neural stem cell mitosis that are reduced during hypothyroidism, did not differ between genotypes. However, in TTR null mice the level of apoptosis, the fate of most progenitor cells, was as low as that in brains of hypothyroid wildtype mice. Thus, lack of TTR results in reduced availability of TH to progenitor cells in the SVZ. We show that proliferation and apoptosis in the SVZ neural stem cell niche are differentially affected by the lack of TTR synthesis.

Cardiac and clinical phenotype in Barth syndrome.

OBJECTIVE: Barth syndrome, an X-linked disorder that is characterized by cardiomyopathy, neutropenia, skeletal myopathy, and growth delay, is caused by mutations in the taffazin gene at Xq28 that result in cardiolipin deficiency and abnormal mitochondria. The clinical phenotype in Barth syndrome has not been characterized systematically, and the condition may be underrecognized. We sought to evaluate extent of cardioskeletal myopathy, potential for arrhythmia, delays in growth, and biochemical correlates of disease severity in patients with this disorder. METHODS: We conducted an observational, cross-sectional study of the largest cohort of patients with Barth syndrome to date (n = 34; age range: 1.2-22.6 years). Evaluation included echocardiography, electrocardiography (standard and signal-averaged), microvolt T wave alternans analysis, biochemical and hematologic laboratory analyses, and physical therapy evaluation of skeletal myopathy. RESULTS: Family history was positive for confirmed or suspected Barth syndrome in 63%. Ninety percent of patients had a clinical history of cardiomyopathy (mean age at diagnosis of cardiomyopathy: 5.5 months; at genetic confirmation of Barth syndrome: 4.6 years). Echocardiography revealed a mean ejection fraction of 50% +/- 10%, mean fractional shortening of 28% +/- 5%, and mean left ventricular end-diastolic volume z score of 1.9 +/- 1.8. Left ventricular morphology demonstrated increased trabeculations or true noncompaction in 53%. Of 16 patients who were evaluated at > or = 11 years of age, 7 (43%) had documented ventricular arrhythmia. Growth deficiency was present (mean weight percentile: 15%; mean height percentile: 8%). Laboratory analysis revealed low total white blood cell count (absolute count: < 4000 cells per microL) in 25% of those who were not on granulocyte colony-stimulating factor. Hypocholesterolemia was present in 24%, decreased low-density lipoprotein cholesterol in 56%, low prealbumin in 79%, and mildly elevated creatine kinase in 15%. CONCLUSIONS: Our cohort demonstrated clinical variability, but most had cardiomyopathy and diminished growth velocity, with a propensity toward neutropenia and low cholesterol. There was increased incidence of ventricular arrhythmia, predominantly in adolescents and young adults. Barth syndrome should be considered when boys present with cardiomyopathy, especially when associated with increased left ventricular trabeculations, neutropenia, skeletal muscle weakness, or family history indicating an X-linked pattern of inheritance.

Doxycycline disrupts transthyretin amyloid: evidence from studies in a FAP transgenic mice model.

Familial amyloidotic polyneuropathy is an autosomal dominant disorder mainly characterized by the extracellular deposition of transthyretin, with special involvement of the peripheral nerve. Several animal models have been generated, including transgenic mice carrying the most prevalent TTR mutation (TTR Val30Met). TTR-Val30Met mice without endogenous TTR (TTR-Val30Met X TTR-KO) were previously analyzed in our laboratory and approximately 60% of the animals over 1 year of age were found to have deposition as amyloid, i.e., with Congo red (CR) -positive material, constituting a good tool to investigate the effect of drugs on TTR deposition and fibrillogenesis. We recently showed that the drug doxycycline acts in vitro as a TTR fibril disrupter. In the present work we assessed the activity of this drug in vivo in the TTR-Met30Val X TTR-KO mice. Doxycycline was administrated in the drinking water to 23- to 28-month-old mice over a period of 3 months. Immunohistochemistry analyses revealed no differences in nonfibrillar TTR deposition between treated (n=11) and untreated mice (n=11). However, CR-positive material was observed only in the control group (untreated) whereas none of the animals treated with doxycycline was CR-positive. Immunohistochemistry for several markers associated with amyloid, such as matrix metalloproteinase-9 (MMP-9) and serum amyloid P component (SAP), was performed. MMP-9 was altered with significantly lower levels in treated animals compared with the control group. Mouse SAP was absent in treated animals, being observed only in untreated animals presenting TTR congophilic deposits. These results indicate that doxycycline is capable of disrupting TTR CR-positive amyloid deposits and decreases standard markers associated with fibrillar deposition, being a potential drug in the treatment of amyloidosis.

Transthyretin knockouts are a new mouse model for increased neuropeptide Y.

Transthyretin (TTR) has access to the brain and nerve through the blood and cerebrospinal fluid. To investigate TTR function in nervous system homeostasis, differential gene expression in wild-type (WT) and TTR knockout (KO) mice was assessed. Peptidylglycine alpha-amidating monooxygenase (PAM), the rate-limiting enzyme in neuropeptide maturation, is overexpressed in the peripheral (PNS) and central nervous system (CNS) of TTR KOs that, consequently, display increased neuropeptide Y (NPY) levels. NPY acts on energy homeostasis by increasing white adipose tissue lipoprotein lipase (LPL) and decreasing thermogenesis; accordingly, we show increased LPL expression and activity in white adipose tissue, PNS, and CNS as well as decreased body temperature in TTR KOs. Associated to increased NPY levels, TTR KOs display increased carbohydrate consumption and preference. In neuronal cells, absence of TTR is related to increased PAM activity, NPY levels and LPL expression, reinforcing that TTR is involved in neuropeptide maturation and that increased NPY correlates with LPL overexpression in the nervous system. Furthermore, we provide molecular insights to the reduced depressive behavior of TTR KOs, as NPY is anti-depressant. Our findings demonstrate that TTR KOs are a model for increased NPY and that TTR plays a role in nervous system physiology.