Proteomics analysis reveals the effect of 1α,25(OH)2VD3-glycosides on development of early testes in piglets.

1α,25(OH)2VD3 is the most active form of VD3 in animals. It plays an important role in regulating mineral metabolism but also in reproduction. Testes are the main reproductive organs of male mammals. Our research aims to reveal the effect of 1α,25(OH)2VD3-glycosides on development of early testes in piglets. 140 weaned 21-day old piglets were selected. The piglets were randomly divided into four groups and were fed a commercial diet supplemented with 0, 1, 2 and 4 µg/kg of 1α,25(OH)2VD3, provided as 1α,25(OH)2VD3-glycosides. Sixty days after the start of the experiment, at piglet age 82 days, testes were harvested. The morphology and histology of early testicular development were assessed. In addition, the proteomic TMT/iTRAQ labelling technique was used to analyse the protein profile of the testes in each group. Western blotting was applied to verify the target of differentially abundant proteins (DAPs). The analysis of morphology and histology of testes showed that a certain concentration of 1α,25(OH)2VD3-glycosides had a positive and significant effect on testicular development. And the results of proteomics analysis showed that of the identified 132,715 peptides, 122,755 were unique peptides. 7852 proteins, of which 6573 proteins contain quantitative information. Screening for DAPs focused on proteins closely related to the regulation of testicular development such as steroid hormone synthesis, steroid biosynthesis, peroxisome and fatty acid metabolism pathways. These results indicated that 1α,25(OH)2VD3 is involved in the regulation of early testicular development in piglets. At the same time, these findings provide valuable information for the proteins involved in the regulation of testicular development, and help to better understand the mechanisms of 1α,25(OH)2VD3 in regulating the development of piglets' testes.

[Effects of temperature and photoperiod on body mass, energy budget and digestive tract morphology in Pycnonotus sinensis.] / æ¸©åº¦ä¸Žå ‰å‘¨æœŸå¯¹ç™½å¤´é¹Žä½“è´¨é‡ã€èƒ½é‡æ”¶æ”¯å’Œæ¶ˆåŒ–é“å½¢æ€çš„å½±å“.

Temperature and photoperiod are proximate environmental factors that have an important influence on the morphological, physiological and behavioral adjustments animals performance du-ring seasonal acclimatization. In this study, the effects of temperature and photoperiod on phenotypic flexibility in body mass, energy budget and digestive tract morphology in the Chinese bulbul (Pycnonotus sinensis) were examined, and the relationship between energy budget and digestive tract morphology was analyzed. Twelve male and sixteen female Chinese bulbuls were randomly assigned into 4 experimental groups so that each group was comprised of three males and four females. The groups were: 1) a warm and long photoperiod (30 ℃, 16 light: 8 dark) group, 2) a warm and short photoperiod (30 ℃, 8 light: 16 dark) group, 3) a cold and long photoperiod (10 ℃, 16 light:8 dark) group, and 4) a cold and short photoperiod (10 ℃, 8 light: 16 dark) group. Each group was acclimated to its respective temperature and photoperiod for 4 weeks. Birds in the cold temperature and short photoperiod group underwent a significant increase in body mass, gross energy intake (GEI) and digestible energy intake (DEI) compared to the other three groups, and there was a significant interaction between temperature and photoperiod on gross energy intake and diges-tible energy intake. The mass of the stomach, small intestine, rectum, and total digestive tract, all increased significantly in cold temperature treatment groups compared to those acclimated to a relatively warm temperature. There was a significant, positive correlation between GEI and DEI resi-duals and those of the length and dry mass of the small intestine and total digestive tract. These results suggested that the Chinese bulbul met the increased energy demands of winter (colder tempe-ratures and reduced foraging time due to shorter day-length) by increasing its body mass, digestible energy intake and digestive tract size.

Cyanogenic glycosides: synthesis, physiology, and phenotypic plasticity.

Cyanogenic glycosides (CNglcs) are bioactive plant products derived from amino acids. Structurally, these specialized plant compounds are characterized as α-hydroxynitriles (cyanohydrins) that are stabilized by glucosylation. In recent years, improved tools within analytical chemistry have greatly increased the number of known CNglcs by enabling the discovery of less abundant CNglcs formed by additional hydroxylation, glycosylation, and acylation reactions. Cyanogenesis--the release of toxic hydrogen cyanide from endogenous CNglcs--is an effective defense against generalist herbivores but less effective against fungal pathogens. In the course of evolution, CNglcs have acquired additional roles to improve plant plasticity, i.e., establishment, robustness, and viability in response to environmental challenges. CNglc concentration is usually higher in young plants, when nitrogen is in ready supply, or when growth is constrained by nonoptimal growth conditions. Efforts are under way to engineer CNglcs into some crops as a pest control measure, whereas in other crops efforts are directed toward their removal to improve food safety. Given that many food crops are cyanogenic, it is important to understand the molecular mechanisms regulating cyanogenesis so that the impact of future environmental challenges can be anticipated.

Cytotoxic, antioxidant, antimicrobial properties and chemical composition of rose petals.

BACKGROUND: Rosa rugosa petals are used for production of teas, jams, wines and juices. Despite the wide availability of rose cultivars, comprehensive information on petal chemical composition and healthful properties is still lacking. Therefore, the aim of this study was analysis of cytotoxic, antioxidant and antimicrobial activity and chemical composition of rugosa rose petals. RESULTS: Petals of R. rugosa were evaluated for their cytotoxic effect against cervical (HeLa) and breast cancer (T47D) cell lines and for antiradical activity (with DPPH•). As a result, significant cytotoxic (up to 100% of dead cells) and antiradical properties (IC50 1.33-0.08 mg mg⁻¹ DPPH•) were demonstrated. Moreover, notable antimicrobial activity against eight bacterial (i.e. Staphylococcus. epidermidis, S. aureus, Bacillus subtilis, Micrococcus luteus, Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Proteus mirabilis) and two yeast strains (Candida. albicans, C. parapsilosis) was shown. Total phenolic, flavonoid, phenolic acid, tannin, carotenoid and polysaccharide content in petals was determined using spectrophotometric methods. Liquid chromatography-electrospray ionization-tandem mass spectrometry was used to thoroughly analyze phenolic acids and flavonoid glycosides in the methanolic extract and fractions obtained after its separation. Five phenolic acids and six flavonoids previously not reported in the plant material were identified. CONCLUSION: This is the first such detailed report on chemical composition and biological activity of R. rugosa petals.

Application of an efficient strategy for discovery and purification of bioactive compounds from Chinese herbal medicines, a case study on the Puerariae thomsonii Flos.

In this study, an efficient strategy based on bioassay-guided fractionation, high-performance liquid chromatography/electrospray ionization quadrupole time-of-flight mass spectrometry (HPLC-ESI-Q/TOF-MS) and high-speed counter-current chromatography (HSCCC) was established to screen and purify bioactive compounds from Chinese herbal medicines (CHMs). This screening system was efficient and successfully applied to reveal anti-prostate cancer candidates from Puerariae thomsonii Flos. As a result, an active fraction with strong in vitro anti-prostate cancer activity was obtained, and the main compounds in the fraction were purified by HSCCC, giving 82 mg of tectoridin, 36 mg of tectorigenin-7-O-[ß-D-xylopyranosyl-(1→6)-ß-D-glucopyranoside and 64 mg of tectorigenin. Among them, tectorigenin, possessing the highest anti-prostate cancer activity with IC50 value of 0.08 µM, has priority to be lead compound. The results of this work demonstrated that the developed method was efficient and could be employed for the rapid screening, identification and purification of active components from CHMs.

Contrasting dose-effects of multi-glycoside of Tripterygium wilfordii HOOK. f. on glomerular inflammation and hepatic damage in two types of anti-Thy1.1 glomerulonephritis.

Multi-glycoside, one of the extracted compounds from Tripterygium wilfordii HOOK. f. (GTW), has been shown to be clinically effective in suppressing glomerular inflammation in chronic kidney disease. However, its clinical application is often limited by its cytotoxic actions on the liver. This study was performed to contrast the dose-effects of GTW on glomerular inflammation and hepatic damage in two types of anti-Thy1.1 glomerulonephritis (GN). Rats with acute or chronic anti-Thy1.1 GN were either left untreated (the Vehicle group) or treated with a high or low dose of GTW and sacrificed on day 7 or day 45. GTW was administrated 3 days before or at the same time as the antibody injection and lasted until sacrifice. GTW at high dose ameliorated glomerular macrophage accumulation, mesangial proliferation, proteinuria, and interleukin-2 expression in the acute anti-Thy1.1 GN model, but caused structural and functional lesions in the liver. In contrast, GTW at low dose improved activated macrophage and T lymphocyte infiltration, mesangial injury, proteinuria, and interleukin-2 and interferon-γ expressions without hepatic toxicity in the chronic model of GN induced by anti-Thy1.1 antibody. In conclusion, GTW at low dose not only effectively inhibits glomerular inflammation but also avoids severe injuries to the liver.

Glycosides, depression and suicidal behaviour: the role of glycoside-linked proteins.

Nowadays depression and suicide are two of the most important worldwide public health problems. Although their specific molecular mechanisms are still largely unknown, glycosides can play a fundamental role in their pathogenesis. These molecules act presumably through the up-regulation of plasticity-related proteins: probably they can have a presynaptic facilitatory effect, through the activation of several intracellular signaling pathways that include molecules like protein kinase A, Rap-1, cAMP, cADPR and G proteins. These proteins take part in a myriad of brain functions such as cell survival and synaptic plasticity. In depressed suicide victims, it has been found that their activity is strongly decreased, primarily in hippocampus and prefrontal cortex. These studies suggest that glycosides can regulate neuroprotection through Rap-1 and other molecules, and may play a crucial role in the pathophysiology of depression and suicide.

The phytotoxin fusicoccin promotes platelet aggregation via 14-3-3-glycoprotein Ib-IX-V interaction.

The fungal toxin fusicoccin induces plant wilting by affecting ion transport across the plasma membrane of plant cell. The activity of this toxin is so far unknown in humans. In the present study we show that fusicoccin is able to affect the platelet aggregation process. The toxin associates with platelet intracellular binding sites and induces aggregation in platelet-rich plasma in a dose-dependent manner. We identified the adhesion receptor glycoprotein Ib-IX-V as fusicoccin target. The toxin promotes the binding of the regulatory 14-3-3 proteins to glycoprotein Ibα and hampers that to glycoprotein Ibß subunit. As a result, platelet adhesion to von Willebrand factor is stimulated, leading to platelet spreading and integrin αIIbß3 activation. We anticipate the present study to be a starting point for future therapeutic use of fusicoccin in genetic bleeding diseases characterized by qualitative or quantitative abnormalities of the platelet membrane-adhesion receptors. Furthermore, the present study also sets the stage for future work to determine the potential pharmacological application of fusicoccin as a drug directed to other 14-3-3-target complexes.

Synergistic effect of kaempferol glycosides purified from Laurus nobilis and fluoroquinolones on methicillin-resistant Staphylococcus aureus.

In a previous study, we reported that two kaempferol glycosides isolated from Laurus nobilis L., kaempferol-3-O-alpha-L-(2'',4''-di-E-p-coumaroyl)-rhamnoside (C2) and kaempferol-3-O-alpha-L-(2''-E-p-coumaroyl-4''-Z-p-coumaroyl)-rhamnoside (C3), showed strong antibacterial activities against methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci. Thereafter we found that these compounds greatly reduced the minimum inhibitory concentrations (MICs) of some fluoroquinolones in MRSA. In other words, C2 and C3 greatly potentiated anti-MRSA activity of fluoroquinolones. The effect of C2 and C3 with fluoroquinolones was found to be synergistic. The potentiation activity was observed with hydrophilic fluoroquinolones, such as norfloxacin and ciprofloxacin, but not with hydrophobic quinolones. We also found that norfloxacin reduced MICs of C2 and C3. The effect was synergistic. Possible mechanism of the synergistic effect was discussed.

Sweet antibiotics - the role of glycosidic residues in antibiotic and antitumor activity and their randomization.

A large number of antibiotics are glycosides. In numerous cases the glycosidic residues are crucial to their activity; sometimes, glycosylation only improves their pharmacokinetic parameters. Recent developments in molecular glycobiology have improved our understanding of aglycone vs. glycoside activities and made it possible to develop new, more active or more effective glycodrugs based on these findings - a very illustrative recent example is vancomycin. The majority of attention has been devoted to glycosidic antibiotics including their past, present, and probably future position in antimicrobial therapy. The role of the glycosidic residue in the biological activity of glycosidic antibiotics, and the attendant targeting and antibiotic selectivity mediated by glycone and aglycone in antibiotics some antitumor agents is discussed here in detail. Chemical and enzymatic modifications of aglycones in antibiotics, including their synthesis, are demonstrated on various examples, with particular emphasis on the role of specific and mutant glycosyltransferases and glycorandomization in the preparation of these compounds. The last section of this review describes and explains the interactions of the glycone moiety of the antibiotics with DNA and especially the design and structure-activity relationship of glycosidic antibiotics, including their classification based on their aglycone and glycosidic moiety. The new enzymatic methodology 'glycorandomization' enabled the preparation of glycoside libraries and opened up new ways to prepare optimized or entirely novel glycoside antibiotics.

TNF-alpha induction of GM2 expression on renal cell carcinomas promotes T cell dysfunction.

Previous studies from our laboratory demonstrated the role of tumor-derived gangliosides as important mediators of T cell apoptosis, and hence, as one mechanism by which tumors evade immune destruction. In this study, we report that TNF-alpha secreted by infiltrating inflammatory cells and/or genetically modified tumors augments tumor-associated GM2 levels, which leads to T cell death and immune dysfunction. The conversion of weakly apoptogenic renal cell carcinoma (RCC) clones to lines that can induce T cell death requires 3-5 days of TNF-alpha pretreatment, a time frame paralleling that needed for TNF-alpha to stimulate GM2 accumulation by SK-RC-45, SK-RC-54, and SK-RC-13. RCC tumor cell lines permanently transfected with the TNF-alpha transgene are similarly toxic for T lymphocytes, which correlates with their constitutively elevated levels of GM2. TNF-alpha increases GM2 ganglioside expression by enhancing the mRNA levels encoding its synthetic enzyme, GM2 synthase, as demonstrated by both RT-PCR and Southern analysis. The contribution of GM2 gangliosides to tumor-induced T cell death was supported by the finding that anti-GM2 Abs significantly blocked T cell apoptosis mediated by TNF-alpha-treated tumor cells, and by the observation that small interfering RNA directed against TNF-alpha abrogated GM2 synthase expression by TNF-transfected SK-RC-45, diminished its GM2 accumulation, and inhibited its apoptogenicity for T lymphocytes. Our results indicate that TNF-alpha signaling promotes RCC-induced killing of T cells by stimulating the acquisition of a distinct ganglioside assembly in RCC tumor cells.

A male gift to its partner? Cyanogenic glycosides in the spermatophore of longwing butterflies (Heliconius).

Males of several insect species transfer nuptial gifts to females during mating, typically in the form of a protein-rich spermatophore. In chemically defended species, males could potentially enhance such a gift with chemicals that help protect the female, her eggs, or both. This was shown for lepidopteran species that accumulate pyrrolizidine alkaloids. Most Heliconius butterflies are presumably protected from predators by virtue of de novo synthesized and/or sequestered cyanogenic glycosides. Males of Heliconius species are known to transfer nutritional gifts to the females but whether defensive chemicals could also be transferred is not known. To ascertain whether transfer of cyanogens occurs, we dissected freshly mated females from nine different Heliconius species and analyzed spermatophores for cyanogenic glycosides. We found cyanogens in the spermatophores of all nine species. This is the first time cyanogenic glycosides are reported in the spermatophores of arthropods. We discuss the implications of these findings for Heliconius biology and for other cyanogenic insects as well. We suggest that chemically defended species commonly lace their nuptial gifts with defensive chemicals to improve gift quality.

The alpha-(1-->6) glycosidic linkage as a novel conformational entropic regulator in osmoregulated periplasmic alpha-cyclosophorohexadecaose.

Molecular dynamics simulations were performed to explain the conformational effect of an alpha-(1-->6)-glycosidic linkage upon the cyclic osmoregulated periplasmic glucan (OPG) produced by Xanthomonas campestris pv. citri. We suggest that a single alpha-(1-->6)-glycosidic linkage in cyclic OPG functions as a novel entropic regulator, which reduces the conformational entropy of cyclic OPG and increases the motional entropy of solvent water molecules.

De novo regeneration of Scrophularia yoshimurae Yamazaki (Scrophulariaceae) and quantitative analysis of harpagoside, an iridoid glucoside, formed in aerial and underground parts of in vitro propagated and wild plants by HPLC.

A protocol for de novo regeneration and rapid micropropagation of Scrophularia yoshimurae (Scrophulariaceae) has been developed. Multiple shoot development was achieved by culturing the shoot-tip, leaf-base, stem-node and stem-internode explants on Murashige and Skoog (MS) medium supplemented with 4.44 microM N6-benzyladenine (BA) and 1.07 microM alpha-naphthaleneacetic acid (NAA). Stem-node and shoot-tip explants showed the highest response (100%) followed by stem-internode (74.4%) and leaf-base (7.7%) explants. The shoots were multiplied by subculturing on the same medium used for shoot induction. Shoots were rooted on growth regulator-free MS basal medium and the plantlets were transplanted to soil and acclimatized in the growth chamber. The content of harpagoside, a quantitatively predominant iridoid glycoside, in different plant material was determined by high performance liquid chromatography (HPLC). The analysis revealed that the content of harpagoside in the aerial and underground parts of S. yoshimurae was significantly higher than the marketed crude drug (underground parts of Scrophularia ningpoensis).

Glycosides in medicine: "The role of glycosidic residue in biological activity".

Numbers of biologically active compounds are glycosides. Sometimes, the glycosidic residue is crucial for their activity, in other cases glycosylation only improves pharmacokinetic parameters. Recent developments in molecular glycobiology brought better understanding to the aglycone vs. glycoside activities, and made possible to develop new, more active or more effective glycodrugs based on these findings - very illustrative recent example is the story of vancomycin. This paper deals with an array of glycosidic compounds currently used in medicine but also with biological activity of some glycosidic metabolites of the known drugs. It involves glycosides of vitamins, polyphenolic glycosides (flavonoids), alkaloid glycosides, glycosides in the group of antibiotics, glycopeptides, cardiac glycosides, steroid and terpenoid glycosides etc. The physiological role of the glycosyl and structure-activity relations (SAR) in the glycosidic moiety (-ies) are discussed.

Host recognition by the tobacco hornworm is mediated by a host plant compound.

It is generally believed that animals make decisions about the selection of mates, kin or food on the basis of pre-constructed recognition templates. These templates can be innate or acquired through experience. An example of an acquired template is the feeding preference exhibited by larvae of the moth, Manduca sexta. Naive hatchlings will feed and grow successfully on many different plants or artificial diets, but once they have fed on a natural host they become specialist feeders. Here we show that the induced feeding preference of M. sexta involves the formation of a template to a steroidal glycoside, indioside D, that is present in solanaceous foliage. This compound is both necessary and sufficient to maintain the induced feeding preference. The induction of host plant specificity is at least partly due to a tuning of taste receptors to indioside D. The taste receptors of larvae fed on host plants show an enhanced response to indioside D as compared with other plant compounds tested.

Brain "ouabain" and angiotensin II contribute to cardiac dysfunction after myocardial infarction.

In chronic heart failure (CHF), sympathetic activity increases in parallel with the impairment of left ventricle (LV) function, and sympathetic hyperactivity has been postulated to contribute to the progression of heart failure. In the brain, compounds with ouabain-like activity ("ouabain," for brevity) and the renin-angiotensin system contribute to sympathetic hyperactivity in rats with CHF after myocardial infarction (MI). In the present studies, we assessed whether, in rats, chronic blockade of brain "ouabain" or the brain renin-angiotensin system inhibits the post-MI LV dysfunction. In rats, an MI was induced by acute coronary artery ligation. At either 0.5 or 4 wk post-MI, chronic treatment with Fab fragments for blocking brain "ouabain" or with losartan for blocking brain AT(1) receptors was started and continued until 8 wk post-MI using osmotic minipumps connected to intracerebroventricular cannulas. At 8 wk post-MI, in conscious rats, LV pressures were measured at rest and in response to volume and pressure overload, followed by LV passive pressure-volume curves in vitro. At 8 wk post-MI, control MI rats exhibited clear increases in LV end-diastolic pressure (LVEDP) at rest and in response to pressure and volume overload. LV pressure-volume curves in vitro showed a marked shift to the right. Intravenous administration of the Fab fragments or losartan at rates used for central blockade did not affect these parameters. In contrast, chronic central blockade with either Fab fragments or losartan significantly lowered LVEDP at rest (only in 0.5- to 8-wk groups) and particularly in response to pressure or volume overload. LV dilation, as assessed from LV pressure-volume curves, was also significantly inhibited. These results indicate that chronic blockade of brain "ouabain" or brain AT(1) receptors substantially inhibits development of LV dilation and dysfunction in rats post-MI.

Na+,K+-ATPase inhibitors in rat urine: origins and physiological significance.

To identify the origins and structures of mammalian tissue-derived Na+,K+-ATPase inhibitors, we investigated the tissue distribution of inhibitors in rats. Among many tissues tested, urine was found to contain high levels of many inhibitors. The structures of the two major inhibitors were identified as neoconvalloside and periplogenin monorhamnoside, which are derivatives of strophanthidin. Urinary levels of these inhibitors, however, decreased considerably after changing the diet from the regular diet to purified synthetic diet, suggesting that the majority of the urinary inhibitors are of dietary origin. Investigation of the ingredients of the diet further revealed that alfalfa meal and ground oats are the major sources of these cardiac glycosides. As to the physiological relevance of the cardiac glycosides, a low concentration (1-50 nM) of ouabain dose-dependently enhanced aldosterone secretion from adrenal glomerulosa cells by an increase in local renin release. Ouabain was also found to be involved in AT2 receptor-specific expression in rat PC12W cells through an increment in intracellular Na+. These results suggest that Na+,K+-ATPase inhibitors, regardless of the source, are involved in the regulation of blood pressure.

A contribution to the regulation of proteoglycan production: modulation by TGF alpha, TGF beta and IL-1 of glycosaminoglycan biosynthesis on beta-D-xyloside in chick embryo fibroblasts.

In order to elucidate the mechanisms determining the variability in the proteoglycan structure and the factors involved in this determination, we treated chick embryo skin fibroblasts with beta-D-xyloside to obtain glycosaminoglycan chains deprived of core proteins, and with different cytokines (transforming growth factor alpha and beta, interleukin-1) to produce variability. The different cytokines specifically regulate both cellular and extracellular amount and composition of glycosaminoglycans. Beta-D-xyloside treatment does not change protein content and protein synthesis, whereas it increases overall extracellular sulphated glycosaminoglycan production, heparan sulphate and chondroitin sulphate content, and reduces that of dermatan sulphate. This indicates that the core protein regulates quantitative proteoglycan production, and probably directs (with appropriate signals) the core oligosaccharide bound to it to the right synthesizing enzymes. The modulatory action of the different cytokines on sulphated glycosaminoglycan production and classes remains, even though the core protein is absent. This indicates that the cytokines also act on the glycosyltransferases. Our results suggest that the proteoglycan production may be subject to a double control, one of which is at the level of the core protein and the other, mediated by environmental signals, at the level of glycosaminoglycan synthesizing enzymes.

Glycosyl phosphatidylinositol in thyroid: cell signalling or protein anchor?

During the last 10 years, attention has been focused on the stimulation by various agonists of the hydrolysis of phosphatidylinositol bis-phosphate into the second messengers inositol tris-phosphate and diacylglycerol. Two other aspects of the metabolism of phosphoinositides were therefore not paid sufficient attention. The first one was the release by insulin of a glycosyl inositol-phosphate from a glycosyl phosphatidylinositol, the hydrosoluble product being able to reproduce some of the hormone effects; the second was the discovery that several membrane proteins were anchored via a glycosyl phosphatidylinositol. For over 20 years, we have been interested in the effect of thyreostimulin (TSH) on the turnover of phosphatidylinositol in pig thyrocyte. As this effect did not seem to result from the hydrolysis of phosphatidylinositol bis-phosphate we explored another possibility, the synthesis of glycosyl inositol-phosphate. We have shown that, in cultured pig thyrocytes, TSH stimulates the release of the polar head of a glycosyl phosphatidylinositol. This soluble glycosyl inositol-phosphate which acts as insulin on adipocyte, modulates the cAMP accumulation and iodine metabolism in thyrocytes and could be held responsible for the cAMP independent effects of TSH. However, we do not yet know if there is a link between the glycosyl phosphatidylinositol sensitive to TSH and the anchor membrane protein. To date, the amount of 2 of these proteins: NAD glyco-hydrolase in thyroid cell membranes and heparan sulfate proteoglycan have been shown to be increased by TSH treatment of thyroid cells.