A novel study approach on Scutellaria altissima L. cultivated at the Ghirardi Botanic Garden (Lombardy, Italy).

Within an Open Science project, research was carried out to describe to the public of the Ghirardi Botanic Garden (BS, Lombardy, Italy) the invisible features of plants. This work is dedicated to Scutellaria altissima L. (Lamiaceae). Micromorphological, histochemical and phytochemical investigations were conducted on the vegetative and reproductive organs to correlate the structures involved in the emission of substances and their unique productivity. This work reports volatile organic compound (VOC) profiles of leaves and flowers and the composition of essential oil (EO) obtained from aerial parts of plants cultivated in Italy that have never been described before. Three morphotypes of glandular trichomes were observed: peltate, short-stalked capitate and long-stalked capitate. Peltate trichomes were the main producers of terpenes, short-stalked capitates of polysaccharides and long-stalked capitates of terpenes and polyphenols. The leaf VOC profile showed heterogeneous composition, with non-terpene derivatives as the major chemical class (71.04%), while monoterpene hydrocarbons represented almost the totality of the flower (99.73%). The leaf presented a higher number of total (37 versus 11) and exclusive (33 versus 7) compounds. (Z)-3-Hexenol acetate was most abundant in the leaf and (E)-ß-ocimene in the flower. Four common compounds were detected: ß-pinene, ß-caryophyllene, γ-muurolene and germacrene-D. The EO contaied 21 compounds, dominated by ß-caryophyllene, linalool and hexahydrofarnesyl acetone. This research allowed us to correlate morphotypes of the secretory structures with the production of secondary metabolites, with the aim of providing the public of the Ghirardi Botanic Garden with a dedicated iconographic approach, which accounts for olfactory perception linked to S. altissima.

Phosphorylation of SOS1 on tyrosine 1196 promotes its RAC GEF activity and contributes to BCR-ABL leukemogenesis.

Son of Sevenless 1 (SOS1) is a dual guanine nucleotide exchange factor (GEF) that activates the small GTPases RAC and RAS. Although the molecular mechanisms of RAS GEF catalysis have been unveiled, how SOS1 acquires RAC GEF activity and what is the physio-pathological relevance of this activity is much less understood. Here we show that SOS1 is tyrosine phosphorylated on Y1196 by ABL. Phosphorylation of Y1196 controls SOS1 inter-molecular interaction, is required to promote the exchange of nucleotides on RAC in vitro and for platelet-derived growth factor (PDGF) activation of RAC- and RAC-dependent actin remodeling and cell migration. SOS1 is also phosphorylated on Y1196 by BCR-ABL in chronic myelogenous leukemic cells. Importantly, in these cells, SOS1 is required for BCR-ABL-mediated activation of RAC, cell proliferation and transformation in vitro and in a xenograft mouse model. Finally, genetic removal of Sos1 in the bone marrow-derived cells (BMDCs) from Sos1fl/fl mice and infected with BCR-ABL causes a significant delay in the onset of leukemogenesis once BMDCs are injected into recipient, lethally irradiated mice. Thus, SOS1 is required for full transformation and critically contribute to the leukemogenic potential of BCR-ABL.

Hypothyroidism and hyponatremia: data from a series of patients with iatrogenic acute hypothyroidism undergoing radioactive iodine therapy after total thyroidectomy for thyroid cancer.

PURPOSE: The aim of the present study was to evaluate the role of hypothyroidism as a cause of hyponatremia in a clinical model of iatrogenic acute hypothyroidism due to thyroid hormone withdrawal prior to ablative radioactive iodine (RAI) therapy after total thyroidectomy. METHODS: The study group consisted of 101 differentiated thyroid cancer (DTC) patients (77 women and 24 men). Plasma concentration of thyroid-stimulating hormone ([TSH]) and sodium ([Na+]) was evaluated before total thyroidectomy (pre[TSH] and pre[Na+]) and on the day of RAI therapy (post[TSH] and post[Na+]). RESULTS: The frequency of hypothyroidism-associated hyponatremia was 4 % (4/101). Pre[Na+] was significantly higher than post[Na+] (140.7 ± 1.6 vs 138.7 ± 2.3 mEq/L, p = 0.012). Moreover, a linear correlation was identified between pre[Na+] and post[Na+]. CONCLUSIONS: Iatrogenic acute hypothyroidism-related hyponatremia is uncommon. However, because of the significant reduction of [Na+] in the transition from euthyroidism to iatrogenic hypothyroidism, the value of pre[Na+] should be viewed as a parameter to be considered. Since it acts as an independent risk factor for the development of hyponatremia, patients with a pre[Na+] close to the lower limit of normal range may deserve a closer monitoring of [Na+].

Phytochemistry, micromorphology and bioactivities of Ajuga chamaepitys (L.) Schreb. (Lamiaceae, Ajugoideae): Two new harpagide derivatives and an unusual iridoid glycosides pattern.

Ajuga chamaepitys (L.) Schreb, well-known as Camaepitium or Ground Pine, is an annual herb typical of the Mediterranean area accounting several uses in the traditional medicine. In this work we have, analyzed the plant iridoid fraction together with the essential oil composition and study of the plant indumentum. Finally, we assayed the polar extracts and essential oil obtained from the aerial parts for antioxidant activity and cytotoxicity on tumor cells. The analysis of the monoterpene glycosides allowed us to isolate from roots and aerial parts and to structurally elucidate by NMR and MS the following compounds: ajugoside (1), reptoside (2), 8-O-acetylharpagide (3), harpagide (4), 5-O-ß-d-glucopyranosyl-harpagide (5), asperulosidic acid (6), deacetyl asperulosidic acid (7) and 5-O-ß-d-glucopyranosyl-8-O-acetylharpagide (8), among which 5 and 8 were two new natural products. Chemotaxomic relevance of these constituents was discussed. The chemical analysis of A. chamaepitys essential oil by GC-FID and GC-MS showed ethyl linoleate (13.7%), germacrene D (13.4%), kaurene (8.4%), ß-pinene (6.8%), and (E)-phytol (5.3%) as the major volatile components. The micromorphological and histochemical study showed that iridoids and essential oil are mainly produced in the type III capitates and peltate trichomes of leaves and flowers. Biological evaluations of A. chamaepitys polar extracts and essential oil showed that the former were more potent as radical scavengers than the latter. MTT assay revealed that essential oil and ethanolic extracts were moderately cytotoxic on tumor cells with IC50 of 36.88 and 59.24µg/mL on MDA-MB 231 cell line, respectively, and IC50 of 60.48 and 64.12µg/mL on HCT116, respectively.

Neuronal distress induced by low extracellular sodium in vitro is partially reverted by the return to normal sodium.

BACKGROUND: Hyponatremia is associated with negative clinical outcomes even when chronic and mild. It is also known that hyponatremia treatment should be appropriately performed, to avoid dramatic consequences possibly leading to death. We have previously demonstrated that chronically low extracellular [Na(+)], independently of reduced osmolality, is associated with signs of neuronal cell distress, possibly involving oxidative stress. AIM: The aim of the present study was to assess whether the return to normal extracellular [Na(+)] is able to revert neuronal cell damage. METHODS: After exposing SH-SY5Y and SK-N-AS cells to low [Na(+)] and returning to normal [Na(+)], we analyzed cell viability by MTS assay, ROS accumulation by FASCan and expression of anti-apoptotic genes. RESULTS: We found that the viability of cells was restored upon return to normal [Na(+)]. However, when more subtle signs of cell distress were assessed, such as the expression level of the anti-apoptotic genes Bcl-2 and DHCR24 or of the heme oxygenase 1 gene, a complete return to basal values was not observed, in particular in SK-N-AS, even when [Na(+)] was gradually increased. We also demonstrated that the amount of ROS significantly increased in low [Na(+)], thus confirming that oxidative stress appears to contribute to the effects of low [Na(+)] on cell homeostasis. CONCLUSIONS: Overall, this study provided the first demonstration that the correction of chronically low extracellular [Na(+)] may not be able to revert all the cell alterations associated with reduced [Na(+)]. These results suggest that prompt hyponatremia treatment might prevent possible residual abnormalities.

Low extracellular sodium promotes adipogenic commitment of human mesenchymal stromal cells: a novel mechanism for chronic hyponatremia-induced bone loss.

Hyponatremia represents an independent risk factor for osteoporosis and fractures, affecting both bone density and quality. A direct stimulation of bone resorption in the presence of reduced extracellular sodium concentrations ([Na(+)]) has been shown, but the effects of low [Na(+)] on osteoblasts have not been elucidated. We investigated the effects of a chronic reduction of extracellular [Na(+)], independently of osmotic stress, on human mesenchymal stromal cells (hMSC) from bone marrow, the common progenitor for osteoblasts and adipocytes. hMSC adhesion and viability were significantly inhibited by reduced [Na(+)], but their surface antigen profile and immuno-modulatory properties were not altered. In low [Na(+)], hMSC were able to commit toward both the osteogenic and the adipogenic phenotypes, as demonstrated by differentiation markers analysis. However, the dose-dependent increase in the number of adipocytes as a function of reduced [Na(+)] suggested a preferential commitment toward the adipogenic phenotype at the expense of osteogenesis. The amplified inhibitory effect on the expression of osteoblastic markers exerted by adipocytes-derived conditioned media in low [Na(+)] further supported this observation. The analysis of cytoskeleton showed that low [Na(+)] were associated with disruption of tubulin organization in hMSC-derived osteoblasts, thus suggesting a negative effect on bone quality. Finally, hMSC-derived osteoblasts increased their expression of factors stimulating osteoclast recruitment and activity. These findings confirm that hyponatremia should be carefully taken into account because of its negative effects on bone, in addition to the known neurological effects, and indicate for the first time that impaired osteogenesis may be involved.

Management of euvolemic hyponatremia attributed to SIADH in the hospital setting.

Hyponatremia is the most frequent electrolyte disorder in hospitalized patients. Acute and severe hyponatremia can be a life-threatening condition, but recent evidence indicates that also mild and chronic hyponatremia is associated with neurological and extra-neurological signs, such as gait disturbances, attention deficits, falls and fracture occurrence, and bone loss. The syndrome of inappropriate ADH secretion (SIADH) is the most frequent cause of hyponatremia. Hyponatremia secondary to SIADH may result for instance from ectopic release of ADH in lung cancer, from diseases affecting the central nervous system, from pneumonia or other pneumopathies or as a side-effect of various drugs In SIADH, hyponatremia results from a pure disorder of water handling by the kidney, whereas external sodium balance is usually well regulated. Despite increased total body water, only minor changes of urine output and modest oedema are usually seen. Neurological impairment may range from subclinical to life-threatening, depending on the degree and mostly on the rate of serum sodium reduction. The management of hyponatremia secondary to SIADH is largely dependent on the symptomatology of the patient. This review briefly summarizes the main aspects related to hyponatremia and then discusses the available treatment options for the management of SIADH, including vaptans, which are vasopressin receptor antagonists targeted for the correction of euvolemic hyponatremia, such as that observed in SIADH.

Lifestyle and high density lipoprotein cholesterol in postmenopause.

OBJECTIVES: Menopause is characterized by hormonal and metabolic changes. These are linked to increased risk of cardiovascular disease, for which low blood plasma levels of high density lipoprotein (HDL) cholesterol are an independent risk factor. The present study investigated variables linked with basal plasma HDL cholesterol levels and the effects of aerobic training, on their variations, in 40 postmenopausal women. METHODS: We assessed body composition, dietary habits and maximal aerobic capacity of participants. Characteristics of daily physical activity and plasma lipoproteins were measured. The women walked on 4 days/week, for 14 weeks, at moderate intensity, and they were grouped according to the resulting tertiles of basal plasma HDL cholesterol levels. RESULTS: Logistic regression analysis showed that waist-to-hip ratio and number of daily bouts of moderate-intensity physical activity, held for at least 10 consecutive minutes (B10m/day), are predictive variables of basal plasma HDL cholesterol levels. After the training period, the first and second tertiles increased plasma HDL cholesterol levels, while the third tertile decreased plasma HDL cholesterol levels. The tertiles showed different remodelling of spontaneous physical activity: the third tertile reduced B10m/day, while the others did not. CONCLUSIONS: This study provides knowledge about the relationships of plasma HDL cholesterol levels with characteristics of physical activity. Furthermore, it shows that physical exercise engagement can result in negative compensation of spontaneous physical activity that could counteract or reduce the positive effects of the aerobic training on plasma HDL cholesterol levels.

A single-institution restrospective experience of brachytherapy in the treatment of pituitary tumors: transsphenoidal approach combined with (192)Ir-afterloading catheters.

BACKGROUND AND AIM: Radiotherapy may be used as an adjuvant treatment of pituitary adenomas. The aim of our study was to present our experience of multimodal treatment of pituitary adenomas, consisting in temporary implantation of (192)Ir-labeled wires following transphenoidal surgery. SUBJECTS AND METHODS: An observational investigation was performed on a series of 80 patients undergoing surgery (S) for pituitary adenomas between 1982 and 2000, some of whom received post-operative external beam radiotherapy (EBRT) (no.=19 between 1982 and 1990), brachytherapy (B) (no.=35, all after 1991), or both irradiation modalities (EBRT+B) (no.=14). The different treatments were compared in terms of hormonal normalization in the subgroup of patients with hypersecreting adenomas, tumor control, and side effects. RESULTS: Hormonal normalization was obtained in 84% of S+B patients and in 61% of S+EBRT patients. Tumor control was obtained in 74.3% of S+B patients and in 63.1% of S+EBRT patients. Anterior pituitary hormones deficits ranged from 8.6-34% in S+B patients and from 15.8-47.4% in S+EBRT patients, after a mean follow-up of 14 yr. The latter group also showed a higher rate of multiple deficits (42.1% vs 22.8%). Diabetes insipidus and other major complications were rare events in all groups. CONCLUSIONS: We presented one original experience regarding brachytherapy in the management of pituitary tumors, which turned out to be effective and safe. Additional prospective, and possibly randomized, studies should clarify whether in the era of 3-dimensional conformal radiotherapy and stereotactic radiotherapy this treatment modality may still have a role.

Rosiglitazone stimulates adipogenesis and decreases osteoblastogenesis in human mesenchymal stem cells.

Thiazolidinediones (TZD) are widely prescribed for the treatment of Type 2 diabetes. Increased loss of bone mass and a higher incidence of fractures have been associated with the use of this class of drugs in post-menopausal women. In vitro studies performed in rodent cell models indicated that rosiglitazone (RGZ), one of the TZD, inhibited osteoblastogenesis and induced adipogenesis in bone marrow progenitor cells. The objective of the present study was to determine for the first time the RGZ-dependent shift from osteoblastogenesis toward adipogenesis using a human cell model. To this purpose, bone marrow-derived mesenchymal stem cells were characterized and induced to differentiate along osteogenic and adipogenic lineages. We found that the exposure to RGZ potentiated adipogenic differentiation and shifted the differentiation toward an osteogenic phenotype into an adipogenic phenotype, as assessed by the appearance of lipid droplets. Accordingly, RGZ markedly increased the expression of the typical marker of adipogenesis fatty-acid binding protein 4, whereas it reduced the expression of Runx2, a marker of osteoblastogenesis. This is the first demonstration that RGZ counteracts osteoblastogenesis and induces a preferential differentiation into adipocytes in human mesenchymal stem cells.

[Nf-kB transcription factor: role in the pathogenesis of inflammatory, autoimmune, and neoplastic diseases and therapy implications]. / Il fattore di trascrizione NF-kB: ruolo nella patogenesi delle malattie infiammatorie, autoimmuni, neoplastiche e implicazioni terapeutiche.

PURPOSE: Description of the involvement of the transcription factor NF-kB in inflammatory, autoimmune and neoplastic processes. Clinical implications from basic research. DESIGN: Review of the most significant data reported in the literature and personal publications. RESULTS: NF-kB is an ubiquitous transcription factor member of the proto-oncogene family rel. NF-kB regulates the expression of several genes involved in inflammatory and immune responses. The classical activated form of NF-kB consists of the p50/p65 heterodimer, different dimers may be formed with members of rel, AP1, steroid hormones receptors family. Many studies suggest that NF-kB should be considered as an important mechanisms of inflammatory processes and autoimmune diseases. Many important anti-inflammatory drugs and immunosuppressants inhibit NF-kB. Several observations have suggested a role of the inappropriate activation of NF-kB in cell proliferation, transformation, and tumor development, mainly lymphomas. Conversely, it has been proposed that the activation of NF-kB in immune cells may contribute to anti-tumor immunity. CONCLUSIONS: NF-kB is an optimal target of anti-inflammatory and immunosuppresant therapies. Molecular studies on NF-kB are very important to understand the pathogenesis of inflammatory, autoimmune and neoplastic diseases, and to identify new drugs that inhibit NF-kB activation.

Transforming growth factor-beta1 down-regulation of major histocompatibility complex class I in thyrocytes: coordinate regulation of two separate elements by thyroid-specific as well as ubiquitous transcription factors.

Transforming growth factor (TGF)-beta1-decreased major histocompatibility complex (MHC) class I gene expression in thyrocytes is transcriptional; it involves trans factors and cis elements important for hormone- as well as iodide-regulated thyroid growth and function. Thus, in rat FRTL-5 thyrocytes, TGF-beta1 regulates two elements within -203 bp of the transcription start site of the MHC class I 5'-flanking region: Enhancer A, -180 to -170 bp, and a downstream regulatory element (DRE), -127 to -90 bp, that contains a cAMP response element (CRE)-like sequence. TGF-beta1 reduces the interaction of a NF-kappaB p50/fra-2 heterodimer (MOD-1) with Enhancer A while increasing its interaction with a NF-kappaB p50/p65 heterodimer. Both reduced MOD-1 and increased p50/p65 suppresses class I expression. Decreased MOD-1 and increased p50/p65 have been separately associated with the ability of autoregulatory (high) concentrations of iodide to suppress thyrocyte growth and function, as well as MHC class I expression. TGF-beta1 has two effects on the downstream regulatory element (DRE). It increases DRE binding of a ubiquitously expressed Y-box protein, termed TSEP-1 (TSHR suppressor element binding protein-1) in rat thyroid cells; TSEP-1 has been shown separately to be an important suppressor of the TSH receptor (TSHR) in addition to MHC class I and class II expression. It also decreases the binding of a thyroid-specific trans factor, thyroid transcription factor-1 (TTF-1), to the DRE, reflecting the ability of TGF-beta1 to decrease TTF-1 RNA levels. TGF-beta1-decreased TTF-1 expression accounts in part for TGF-beta1-decreased thyroid growth and function, since decreased TTF-1 has been shown to decrease thyroglobulin, thyroperoxidase, sodium iodide symporter, and TSHR gene expression, coincident with decreased MHC class I. Finally, we show that TGF-beta1 increases c-jun RNA levels and induces the formation of new complexes involving c-jun, fra-2, ATF-1, and c-fos, which react with Enhancer A and the DRE. TGF-beta1 effects on c-jun may be a pivotal fulcrum in the hitherto unrecognized coordinate regulation of Enhancer A and the DRE.

Thymosin-alpha1 regulates MHC class I expression in FRTL-5 cells at transcriptional level.

In this study we examined the effect of the synthetic peptide thymosin-alpha1 (T(alpha)1) on MHC class I expression in FRTL-5 cells. Treatment with T(alpha)1 increased expression of MHC class I surface molecules and mRNA, which reached its peak (153 +/- 8 % of the control value) after 12 h. Chloramphenicol acetyltransferase (CAT) analysis, following transfection with a plasmid containing the regulatory sequence of MHC class I (or its deletion derivatives) with the CAT reporter gene, and electrophoretic mobility shift assay experiments demonstrated that the action of T(alpha)1 was at the transcriptional level, and its mechanism of action is likely due to increased binding between the complex p50/fra-2 and the enhancer A sequence of the 5' flanking region of a swine class I gene (PD1). An increase in the expression of MHC class I surface molecules was also observed by flow cytometry in murine and human tumor cell lines and in primary cultures of human macrophages. This study shows for the first time an effect of Talpha1 on the regulation of gene expression at the molecular level, and may further contribute to explaining the results obtained using Talpha1 in the control of infectious diseases and tumor growth.

[The clinical use of human recombinant TSH]. / Impiego clinico del TSH umano ricombinante.

The recent cloning of human TSH-beta gene has allowed the production of recombinant human TSH (rhTSH) by recombinant DNA technology in mammalian cells (Chinese hamster ovary cells). Studies aimed at biochemical and biological characterization have shown that rhTSH, unlike pituitary TSH, is highly sialylated and is biological active in stimulating c-AMP accumulation in FRTL-5 cells. Phase I/II and phase III clinical studies have been performed to evaluate the safety and efficacy of rhTSH in stimulating radioactive iodine uptake in patients after total thyroidectomy for differentiated thyroid cancer. In these patients therapy with thyroid hormones is performed to replace hormone production and to suppress TSH-stimulated tumor growth. To detect residual or recurrent cancer, the therapy has to be withdrawn in order to obtain rise in endogenous TSH to perform a total body scan. rhTSH, as a source of exogenous human TSH, has been shown as an additional diagnostic tool in the follow-up of patients with thyroid cancer. Used in patients maintained on thyroid hormone suppressive therapy, rhTSH enhances the sensitivity of serum Tg testing. Although the sensitivity of scans obtained after rhTSH administration is slightly lower than that after thyroid hormone withdrawal, the use of rhTSH avoids the clinical signs and symptom of hypothyroidism and can be used in selected patients.

Iodide suppression of major histocompatibility class I gene expression in thyroid cells involves enhancer A and the transcription factor NF-kappa B.

High concentrations of iodide can induce transient, clinical improvement in patients with autoimmune Graves' disease. Previous work has related this iodide action to the autoregulatory effect of iodide on the growth and function of the thyroid; more recently, we additionally related this to the ability of iodide to suppress major histocompatibility (MHC) class I RNA levels and antigen expression on thyrocytes. In this report, we describe a transcriptional mechanism involved in iodide suppression of class I gene expression, which is potentially relevant to the autoregulatory action of iodide. Transfection experiments in FRTL-5 cells show that iodide decreases class I promoter activity and that this effect can be ascribed to the ability of iodide to modulate the formation of two specific protein/DNA complexes with enhancer A, -180 to -170 bp, of the class 1 5'-flanking region. Thus, iodide decreases the formation of Mod-1, an enhancer A complex involving the p50 subunit of NF-kappa B and a c-fos family member, fra-2, which was previously shown to be important in the suppression of class I levels by hydrocortisone. Unlike hydrocortisone, iodide also increases the formation of a complex with enhancer A, which we show, in antibody shift experiments, is a heterodimer of the p50 and p65 subunits of NF-kappa B. The changes in these complexes are not duplicated by chloride and are related to the action of iodide on class I RNA levels by the following observations. First, FRTL-5 thyroid cells with an aged phenotype coincidentally lose the ability of iodide to decrease MHC class I RNA levels and to induce changes in either complex. Second, the effect of iodide on class I RNA levels and on enhancer A complex formation with Mod-1 and the p50/p65 heterodimer is inhibited by agents that block the inositol phosphate, Ca++, phospholipase A2, arachidonate signal transduction pathway: acetylsalicylate, indomethacin, and 5,8,11,14-eicosatetraynoic acid. Interestingly, iodide can also decrease formation of the Mod-1 complex and increase formation of the complex with the p50/p65 subunits of NF-kappa B when the NF-kappa B enhancer sequence from the Ig kappa light chain, rather than enhancer A, is used as probe; and both actions mimic the action of a phorbol ester. This suggests that iodide may regulate complex formation with NF-kappa B regulatory elements on multiple genes associated with growth and function, providing a potential mechanism relating the autoregulatory action of iodide on thyroid cells and its action on class I gene expression.

Major histocompatibility class II HLA-DR alpha gene expression in thyrocytes: counter regulation by the class II transactivator and the thyroid Y box protein.

Aberrant expression of major histocompatibility complex (MHC) class II proteins on thyrocytes, which is associated with autoimmune thyroid disease, is mimicked by gamma-interferon (gamma-IFN). To define elements and factors that regulate class II gene expression in thyrocytes and that might be involved in aberrant expression, we have studied gamma-IFN-induced HLA-DR alpha gene expression in rat FRTL-5 thyroid cells. The present report shows that class II expression in FRTL-5 thyrocytes is positively regulated by the class II transactivator (CIITA), and that CIITA mimics the action of gamma-IFN. Thus, as is the case for gamma-IFN, several distinct and highly conserved elements on the 5'-flanking region of the HLA-DR alpha gene, the S, X1, X2, and Y boxes between -137 to -65 bp, are required for class II gene expression induced by pCIITA transfection in FRTL-5 thyroid cells. CIITA and gamma-IFN do not cause additive increases in HLA-DR alpha gene expression in FRTL-5 cells, consistent with the possibility that CIITA is an intermediate factor in the gamma-IFN pathway to increased class II gene expression. Additionally, gamma-IFN treatment of FRTL-5 cells induces an endogenous CIITA transcript; pCIITA transfection mimics the ability of gamma-IFN treatment of FRTL-5 thyroid cells to increase the formation of a specific and novel protein/DNA complex containing CBP, a coactivator of CRE binding proteins important for cAMP-induced gene expression; and the action of both gamma-IFN and CIITA to increase class II gene expression and increase complex formation is reduced by cotransfection of a thyroid Y box protein, which suppresses MHC class I gene expression in FRTL-5 thyroid cells and is a homolog of human YB-1, which suppresses MHC class II expression in human glioma cells. We conclude that CIITA and TSH receptor suppressor element binding protein-1 are components of the gamma-IFN-regulated transduction system which, respectively, increase or decrease class II gene expression in thyrocytes and may, therefore, be involved in aberrant class II expression associated with autoimmune thyroid disease.

Regulation of major histocompatibility complex class I gene expression in thyroid cells. Role of the cAMP response element-like sequence.

The major histocompatibility complex (MHC) class I gene cAMP response element (CRE)-like site, -107 to -100 base pairs, is a critical component of a previously unrecognized silencer, -127 to -90 bp, important for thyrotropin (TSH)/cAMP-mediated repression in thyrocytes. TSH/cAMP induced-silencer activity is associated with the formation of novel complexes with the 38-base pair silencer, whose appearance requires the CRE and involves ubiquitous and thyroid-specific proteins as follows: the CRE-binding protein, a Y-box protein termed thyrotropin receptor (TSHR) suppressor element protein-1 (TSEP-1); thyroid transcription factor-1 (TTF-1); and Pax-8. TTF-1 is an enhancer of class I promoter activity; Pax-8 and TSEP-1 are suppressors. TSH/cAMP decreases TTF-1 complex formation with the silencer, thereby decreasing maximal class I expression; TSH/cAMP enhance TSEP-1 and Pax-8 complex formation in association with their repressive actions. Oligonucleotides that bind TSEP-1, not Pax-8, prevent formation of the TSH/cAMP-induced complexes associated with TSH-induced class I suppression, i.e. TSEP-1 appears to be the dominant repressor factor associated with TSH/cAMP-decreased class I activity and formation of the novel complexes. TSEP-1, TTF-1, and/or Pax-8 are involved in TSH/cAMP-induced negative regulation of the TSH receptor gene in thyrocytes, suppression of MHC class II, and up-regulation of thyroglobulin. TSH/cAMP coordinate regulation of common transcription factors may, therefore, be the basis for self-tolerance and the absence of autoimmunity in the face of TSHR-mediated increases in gene products that are important for thyroid growth and function but are able to act as autoantigens.

Centralized cytogenetic analysis of pediatric acute leukemia: results of an Italian collaborative experience.

BACKGROUND AND OBJECTIVE: Cytogenetic analysis of acute leukemia yields important information which has been demonstrated to be correlated to patient survival. A reference laboratory was created in order to perform karyotype analysis on all cases of acute leukemia enrolled in the AIEOP (Associazione Italiana Emato-Oncologia Pediatrica) protocols. METHODS: From January 1990 to December 1995, 1115 samples of children with ALL or AML were sent in for cytogenetic analysis. The results of cell cultures were screened in the Reference Laboratory and then the fixed metaphases were sent to one of the six cytogenetic laboratories for analysis. RESULTS: The leukemic karyotypes of 556 patients were successfully analyzed. An abnormal clone was detected in 49% of cases of ALL and in 66% of AML. In ALL the most frequent abnormality was 9p rearrangement. Other recurrent abnormalities were t(9;22), t(4;11) and t(1;19). In AML t(8;21), t(15;17) and 11q23 rearrangement were the most frequent structural abnormalities. These findings are similar to the results obtained in other multicenter studies using a similar approach. INTERPRETATION AND CONCLUSIONS: Our data confirm the feasibility of performing cytogenetic analysis in a centralized laboratory on mailed samples of bone marrow and/or peripheral blood; this is very important considering that cytogenetic analysis of neoplastic tissue requires a special laboratory and expert staff.

Purification and characterization of a 90 kDa protein released from human tumors and tumor cell lines.

A novel tumor-associated protein, termed 90K, and recognized by mAb SP-2 was purified from serum of breast cancer patients, ovarian cancer ascitic fluid and conditioned medium of human breast cancer cells. In these three sources, native 90K is present as a high molecular weight complex that was dissociated by SDS-PAGE into a major band of approximately 90,000 Da. On the basis of electrophoretic mobility, buoyant density value, amino acid composition, and immunoreactivity, the 90K from the different sources appeared to be identical. NH2-terminal amino acid sequence revealed no homology to known protein.