Circular RNA circCNOT6L regulates cell development through modulating miR-384/FN1 axis in esophageal squamous cell carcinoma.

OBJECTIVE: In recent years, circular RNAs (circRNAs) and microRNAs (miRNAs) have been shown to be related to the development of esophageal squamous cell carcinoma (ESCC). However, their functional mechanisms remain to be investigated. Herein, we focus our research on the functions and mechanisms of circCNOT6L and miR-384 in ESCC. MATERIALS AND METHODS: The levels of circCNOT6L, miR-384, and fibronectin 1 (FN1) were determined using quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). RNase R was used to investigate circCNOT6L stabilization. Cell proliferation and apoptosis were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry, respectively. Western blot assay was employed to analyze the protein levels of FN1, proliferation-related genes, and iron metabolism-related genes. In addition, the interaction between miR-384 and circCNOT6L or FN1 was predicted by starBase3.0 and confirmed by the Dual-Luciferase reporter assay. Mouse xenograft was carried out to measure the effect of circCNOT6L on tumor growth in vivo. RESULTS: CircCNOT6L and FN1 levels were upregulated, and miR-384 level was downregulated in ESCC tissues/cells. CircCNOT6L knockdown attenuated ESCC cell proliferation and iron metabolism disorder, as well as accelerated apoptosis. Notably, circCNOT6L targeted miR-384, and miR-384 targeted FN1. MiR-384 depletion and FN1 upregulation weakened the effects of circCNOT6L knockdown and miR-384 overexpression on ESCC cell progression, respectively. Besides, circCNOT6L knockdown inhibited tumor growth in vivo. CONCLUSIONS: Our results demonstrated that circCNOT6L positively regulated the development of ESCC cells via modulating miR-384/FN1 axis. Our findings provided a theoretical basis for the therapy of ESCC patients.

Overexpression and amplification of c-myc in the Syrian hamster kidney during estrogen carcinogenesis: a probable critical role in neoplastic transformation.

An estrogen receptor-driven, multistep process for estrogen carcinogenesis in the Syrian hamster kidney is proposed. Because in this species the reproductive and urogenital tracts arise from the same embryonic germinal ridge, it is evident that the kidney has carried over genes that are responsive to estrogens. Using in situ hybridization, overexpression of early estrogen-response genes, i.e., c-myc and c-fos, has been shown to be localized preferentially in early renal tumor foci after 3.5-4.0 months of estrogen treatment. This event coincides with an increased number of S-phase proliferating cell nuclear antigen-labeled cells in these tumor foci, along with a rapid rise in aneuploid frequency in the kidney. Western blot analyses of c-MYC and c-FOS protein products support the overexpression of these genes. Amplification of c-myc, 2.4-3.6-fold, but not of c-fos, was detected in 67% of the primary renal tumors examined, by Southern blot analyses. Consistent chromosomal gains, common to both diethylstilbestrol- and estradiol-induced renal neoplasms, were observed in chromosomes 1, 2, 3, (6), 11, (13), 16, 20, and 21 (chromosome number alterations are indicated in parentheses). Using fluorescence in situ hybridization, the c-myc gene was localized to hamster chromosome 6qb. Chromosome 6 exhibited a high frequency of trisomies and tetrasomies in the kidney after 5.0 months of estrogen treatment and in primary renal tumors. The data presented indicate that estrogen-induced genomic instability may be a key element in carcinogenic processes induced by estrogens.

Expression of the c-jun, jun-B, ets-2 and liver regeneration factor-1 (LRF-1) genes during promotion and progression of rat liver carcinogenesis in the resistant hepatocyte model.

During promotion in the RH-model, the mRNA expression of c-jun and LRF-1 was 2- to 8-fold elevated in both initiated and uninitiated rats receiving 2-AAF. The increase was more pronounced in male than in female rats, and GH treatment of male rats down-regulated the expression towards the level in females. The level in uninitiated 2-AAF-treated livers was as high as in isolated early nodules. jun-B also showed 3- to 8-fold increased expression, but without sex differences. An increased nuclear transcription of the LRF-1 and jun-B genes but not of c-jun was observed. During progression, LRF-1 and ets-2 showed a 2- to 3-fold higher expression in persistent nodules and hepatocellular carcinomas than in the corresponding surrounding liver tissues, whereas the expression of the jun genes was 3- to 4-fold increased both in lesions and in surrounding livers when compared to age-matched control rats. In conclusion, while the changes during promotion might not be connected with control of early focal growth, the increased levels of LRF-1 and ets-2 in advanced lesions might indicate that these genes could contribute to the growth advantage for persistent nodules during progression.

[Effects of morphine of different concentrations on myocardial action potential].

The effects of morphine at different concentrations on myocardial action potential were studied in isolated right ventricular papillary muscles of the guinea pig. It was observed that morphine at low concentrations (0.2-1.6 mumol/L) shortened the action potential duration (ADP) and effective refractory period (ERP) in a concentration dependent manner. These effects could be abolished by naloxone (1 mumol/L), phentolamine, tetraethylammonium (TEA) and cesium chloride (Cs+), but not by verapamil. On the other hand, morphine at high concentrations (15-120 mumol/L) prolonged ADP and ERP in a concentration dependent manner. The effects were unaffected by low dose of naloxone (1.2 mumol/L) but were abolished by high dose of naloxone (10 mumol/L), phentolamine, TEA, Cs+ and verapamil. These results suggest that morphine at low and high concentrations might stimulate different subtypes of opioid receptors. The effects of morphine in low concentrations are associated with the activation of potassium channel, whereas the effects of morphine at high concentrations are associated with the activation of potassium channel, calcium channel or calcium activated potassium channel. The action of opioid receptor was closely related to alpha adrenoreceptors.

Unusual deregulation of cell cycle components in early and frank estrogen-induced renal neoplasias in the Syrian hamster.

There is strong evidence that estrogens are involved in the etiology, promotion and progression of a variety of cancers, including the cancers of the breast and endometrium. The Syrian hamster estrogen-induced, estrogen-dependent renal neoplasm is a well-established animal model used to elucidate the cellular and molecular mechanisms involved in solely estrogen-induced carcinogenic processes. G(1) cell cycle progression was studied in estrogen-induced early renal tumor foci and in large kidney tumors of castrated male hamsters. Levels of cyclin D1, cyclin E and retinoblastoma (pRb) proteins were higher in these renal neoplasias than in adjacent uninvolved renal tissue and kidneys from untreated, age-matched animals. Of particular interest is the presence of a predominant 35 kDa cyclin E protein variant form in primary renal tumors. In addition, amounts of the phosphorylated forms of cyclin-dependent kinases (cdk) 2 and 4 were decreased, and both RNA and protein levels of p27(kip1) (p27), a cyclin-dependent kinase inhibitor, were markedly higher in early and frank renal tumors than in adjacent uninvolved renal tissue and kidneys of untreated, age-matched animals. These changes in cell cycle components coincided with a rise in renal tumor cell proliferation. Binding of the elevated p27 protein to cyclin E, cdk2 and cdk4, however, was not impaired, suggesting that this cell cycle suppressor protein is functional. In addition, cyclin D1-, cdk2-, cdk4- and cyclin E-associated kinase activities were also lower in these estrogen-induced renal neoplasms than in untreated, age-matched kidneys. Interestingly, when compared with untreated kidney tissue, early and frank renal neoplasms had less of the 62 kDa native form of E2F1 and contained a 57 kDa variant form. Thus we have characterized an unusual deregulation of the cell cycle during estrogen-induced renal tumorigenesis in Syrian hamsters which still allows for estrogen-driven kidney tumor cell proliferation and may contribute to the early genomic instability found.

Expression of the liver-enriched transcription factors C/EBP alpha, C/EBP beta, HNF-1, and HNF-4 in preneoplastic nodules and hepatocellular carcinoma in rat liver.

The expression patterns of the liver-enriched transcription factors CCAAT/enhancer-binding protein (C/EBP) alpha and beta and hepatocyte nuclear factor (HNF)-1 and HNF-4 were studied in liver nodules and hepatocellular carcinomas from male rats treated according to the resistant hepatocyte (RH) model. C/EBP alpha expression was lower at the transcriptional, mRNA, and protein levels in persistent nodules than in the respective surrounding livers. Expression was further decreased in the tumors. Transcriptional downregulation of C/EBP alpha gene expression was observed already in very early nodules, isolated 3 wk after partial hepatectomy in the RH model. However, no detectable changes were observed in preneoplastic nodules in the transcription or in steady-state mRNA levels of C/EBP beta, HNF-1, and HNF-4. A slight decrease in C/EBP beta protein and a more pronounced attenuation of HNF-1 and HNF-4 levels was observed in nodules, being 67%, 37%, and 46% of the levels in the corresponding surrounding livers, respectively. In conclusion, differential regulation of several transcription factors that are associated with the maintenance of the differentiated state of the hepatocytes was observed in preneoplastic and neoplastic liver lesions. This could have an impact on the regulation of a wide array of genes during liver carcinogenesis. Furthermore, the attenuation of C/EBP alpha expression, regarded as a negative growth regulator, could contribute to the proliferative advantage of nodules during liver carcinogenesis.

Promotion of estrogen-induced mammary gland carcinogenesis by androgen in the male Noble rat: probable mediation by steroid receptors.

Both endogenous and exogenous estrogen exposure is associated with an increased breast cancer risk. In some studies, elevated serum testosterone levels have also been linked to an increased breast cancer risk. Estrogen alone or combined with progesterone induces high mammary tumor incidences in various strains of both male and female rats. Mammary gland ductal adenocarcinomas were induced after 17beta-estradiol (E2) and testosterone propionate (TP) treatment in male Noble rats. Tumor incidence was 100% after 8-9 months of treatment. Such neoplasms were not detected after either estrogen or androgen exposure alone within this time period. TP alone caused disruption of mammary gland ducts and proliferation of stromal tissue, while E2 treatment alone induced both ductal epithelial growth and nodular atypical hyperplasia. To study the interaction of these hormones in mammary tumorigenesis, sex hormone receptors were characterized in mammary glands of Noble rats. Estrogen receptor-alpha (ER) was detected in age-matched, untreated mammary gland epithelium; in most early atypical hyperplastic lesions appearing after E2 and E2 + TP treatment and in E2 + TP-induced mammary tumors. Two major ER putative isoforms, 116 and 120 kDa, were detected in E2- and E2 + TP-treated mammary glands, and in the induced tumors. A 54 kDa ER protein was found in untreated and TP-treated mammary glands, and in the induced tumors. Both progesterone receptor-B (PR-B) and PR-A2, as well as androgen receptor-B (AR-B) and AR-A isoforms were markedly elevated in all E2 + TP-induced mammary tumors. However, the levels of both PR and AR were very low in mammary glands of E2- and E2 + TP-treated male rats. Low and moderate levels of AR and PR, respectively, were detected in most atypical hyperplastic lesions induced by E2- and E2 + TP-treated mammary glands. These results suggest that androgens may interact with either AR or PR, and perhaps both receptors, in E2 + TP-induced mammary glands and the induced tumors to effect the reduction in latency period, enhance tumor size, and increase incidence to 100%.

Persistent sex differences in growth control of early rat liver lesions are programmed during promotion in the resistant hepatocyte model.

The effects of gonadal hormones on several parameters associated with sex-differentiated promotion in the resistant hepatocyte (RH) model were studied. Male and female rats were initiated with diethylnitrosamine and promoted with 2-acetylaminofluorene (2-AAF) and partial hepatectomy [correction of hepatecomy] (PH). Before promotion, some female rats were ovariectomized, with or without receiving subcutaneous testosterone implants. Rats were killed either at the time of cessation of 2-AAF treatment or 2 weeks later. Ovariectomy decreased the messenger RNA (mRNA) expression of the female-specific cytochrome P450 2C12 (CYP2C12) at the time of PH, but did not increase the male-specific CYP2C11. Testosterone treatment further decreased CYP2C12 and induced CYP2C11 to the level in male liver. Hepatic foci positive for the placental form of glutathione-S-transferase (GST-P) were larger in male than in female rats. Ovariectomy did not affect the size of foci, whereas testosterone treatment increased the size to the male level. At the time of cessation of 2-AAF treatment, the labeling index, determined as cells staining for proliferating cell nuclear antigen, was higher in foci of males and testosterone-treated females than in foci from females with or without ovariectomy, whereas the labeling index in the surrounding hepatocytes was lower in males and testosterone-treated females. Two weeks later, the sex differences in labeling index were still present in foci, but no differences were observed in the surrounding hepatocytes. An elevated c-myc expression was observed in nodules isolated 3 weeks after PH from males and testosterone-treated females, but not in nodules from intact females. In conclusion, ovarian hormones did not affect promotion in the RH-model, whereas testosterone administration to ovariectomized females masculinized growth hormone-regulated hepatic parameters and response to promotion.

A novel hydrogen-bonded microporous framework constructed from two different metal complexes.

A hydrogen-bonded coordination supramolecule, (meso-5,7,7,12,14,14-hexamethyl-1,4,8,11-tetraazacyclotetradecane-kappa4N)nickel(II) [N,N-o-phenylenebis(oxamato)-kappa4O,N,N',O']nickelate(II) dihydrate, [Ni(C16H36N4)][Ni(C10H4N2O6)]*2H2O or [Ni(meso-cth)][Ni(opba)]*2H2O, has been prepared and characterized by X-ray crystallographic analysis. The two complex ions, i.e. [Ni(meso-cth)]2+ and [Ni(opba)]2-, are hydrogen bonded to each other, resulting in two-dimensional neutral supramolecular sheets. The sheets stack along the a direction to produce a three-dimensional architecture with one-dimensional channels in which hydrogen-bonded chains of water molecules are included.

Varying coordination modes and magnetic properties of copper(II) complexes with diazamesocyclic ligands by altering additional donor pendants on 1,5-diazacyclooctane.

A series of new diazamesocyclic ligands based on a diazamesocycle, 1,5-diazacyclooctane (DACO), functionalized by additional donor groups--1,5-bis(N-1-methylimidazol-2-ylmethyl)-1,5- diazacyclooctane (L1), 1-(2-hydroxybenzyl)-1,5-diazacyclooctane (HL2), 1,5-bis(2-hydroxybenzyl)-1,5-diazacyclooctane (H2L3), and 1-(N-1-methylimidazol-2-ylmethyl)-1,5-diazacyclooctane (L4)--and their Cu(II) complexes have been synthesized and characterized. Single-crystal X-ray diffraction analysis of the four Cu(II) complexes revealed that L1 forms a five-coordinate mononuclear complex, HL2 a N3- mu-bridged binuclear complex, H2L3 an oxygen mu-bridged trinuclear complex, and L4 a one-dimensional zigzag coordination polymeric complex with Cu(II). [CuL1ClO4](ClO4) (I): a = 12.194(2) A, b = 13.351(3) A, c = 14.473(3) A, beta = 107.10(3) degrees, Z = 4. [CuL2(N3)]2 (II): a = 8.1864(6) A, b = 18.141(2) A, c = 9.3307(7) A, beta = 103.662(6) degrees, Z = 2. [Cu3(L3)2Cl2] (III): a = 10.7296(13) A, b = 13.7707(17) A, c = 13.5523(17) A, beta = 106.350(3) degrees, Z = 2. ([CuL4Cl]2ClO4) infinity (IV): a = 7.279(1) A, b = 23.695(5) A, c = 19.308(4) A, beta = 100.28(3) degrees, Z = 8. All four complexes crystallize in the monoclinic crystal system with the P2(1)/c space group, and each Cu(II) center coordinated with DACO is pentacoordinated with a distorted square-pyramidal or trigonal-bipyrimidal coordination environment. In complex IV, the binuclear cation unit [CuL4Cl]2(2+) constitutes the fundamental building block of an infinite alternating zigzag chain structure, and the binuclear unit contains two types of geometries around the Cu(II) centers: the Cu(1) center is a distorted square-pyramidal environment, while the Cu(2) is a distorted trigonal-bipyramidal coordination environment. To the best of our knowledge, this is the first Cu(II) complex of a diazamesocyclic ligand with an infinite polymeric structure. The magnetic properties of complexes II, III, and IV have been investigated by variable-temperature magnetic susceptibility measurements in the solid state. The obtained parameters are 2J = 2.06 cm-1 (II), -345.56 cm-1 (III), and -2.60 cm-1 (IV), which differ greatly from ferromagnetic to weak and strong antiferromagnetic coupling. These results unequivocally indicate that the nature of the pendant arms is a key factor governing the structure and properties of the complexes; therefore, the coordination modes and properties of the metal complexes of a diazamesocycle can be controlled by altering the pendant donors on it. Magneto-structural correlation has been precisely analyzed, and the solution properties of these complexes have also been described.

Induction of cathepsin D protein during estrogen carcinogenesis: possible role in estrogen-mediated kidney tubular cell damage.

We have proposed that an early step in estrogen carcinogenesis in the hamster kidney is tubular damage followed by reparative cell proliferation. This tubular injury is progressive and increases in severity with continued estrogen treatment; one pertinent feature is a marked rise in the number of both secondary and tertiary lysosomes. Data presented herein indicate that cathepsin D, an estrogen-responsive lysosomal proteolytic enzyme, is increased in the kidney following estrogen treatment in the hamster. Three isoforms of cathepsin D were detected in estrogen-treated kidneys, 52, 31, and 27 kDa, the major being 52 kDa. At 1 and 3 months of estrogen treatment, 52-kDa cathepsin D content increased 1.4- to 1.6-fold. These changes coincided with a rise in renal estrogen receptor levels during the same estrogen treatment periods. More pronounced rises in cathepsin D levels, 2.7- and 3.5-fold, were seen after 4 and 5 months of estrogen treatment, respectively. A concomitant, 3.0- to 4.0-fold rise in estrogen receptor content was also observed. At 5 months of estradiol or DES treatment, both 27- and 31-kDa isoforms were present in hamster kidneys, in addition to the 52-kDa form. Neither progesterone nor DHT treatment affected the untreated levels of cathepsin D. Interestingly, either concomitant tamoxifen or DHT and estrogen treatment prevented the rise in cathepsin D and estrogen receptor content observed after estrogen treatment alone. Primary estrogen-induced renal tumors and their metastases exhibited markedly elevated levels of all three isoforms of cathepsin D. Immunohistochemical analysis of cathepsin D in kidney sections confirmed the Western blot findings. These data suggest a novel role for estrogen-induced cathepsin D in the hamster kidney during tumorigenesis; that is, mediating renal tubular damage as a prelude to reparative cell proliferation, thus initiating a multi-step estrogen-driven process which leads to renal tumor formation.

Two-dimensional coordination polymers of copper(II) with oxalate: lattice water control of structure.

Three oxalate copper(II) complexes, [Cu(bipy)(C(2)O(4))(H(2)O)].2H(2)O (1), [Cu(nphen)(C(2)O(4))(H(2)O)].2H(2)O (2), and [Cu(phen)(C(2)O(4))(H(2)O)].H(2)O (3) (bipy = 2,2'-bipyridine, nphen = 5-nitro-1,10-phenanthroline and phen = 1,10-phenanthroline), have been synthesized and their crystal structures have been determined. Compound 1 crystallizes in the triclinic space group P1 with a = 7.2554(10) A, b = 10.5712(14) A, c = 10.8178(15) A, alpha = 62.086(2) degrees, beta = 77.478(3) degrees, gamma = 81.773(3) degrees, and Z = 2. Compound 2 crystallizes in the triclinic space group P1 with a = 9.582(2) A, b = 10.086(2) A, c = 10.592(2) A, alpha = 64.18(3) degrees, beta = 79.47(3) degrees, gamma = 60.06(3) degrees, and Z = 2. Compound 3 crystallizes in the monoclinic space group P2(1)/n with a = 8.4655(7) A, b = 9.7057(8) A, c = 17.4572(14) A; beta = 103.865(2) degrees, and Z = 4. The crystal structures of all complexes consist of neutral [Cu(L)(C(2)O(4))(H(2)O)] (L = bipy, nphen, and phen) units and one or two lattice water molecules in the unit cell. Each copper atom in 1, 2, and 3 involves a five-coordinate CuN(2)O(2)O' environment, with a distorted square-pyramidal structure. In 1 and 2, two lattice water molecules are around each unit of [CuL(C(2)O(4))(H(2)O)] (L = bipy and nphen) and form two-dimensional networks. Only one lattice water molecule is found in the unit cell of 3 and the two-dimensional structure is different from 1 and 2. The extended three-dimensional structure is formed through pi-pi interactions between layers. The influences of hydrogen bonds and the sizes and Lewis basicity of ligands to the structures were discussed.

Oxamato-bridged trinuclear Ni(II)Cu(II)Ni(II) complexes with irregular spin state structures and a binuclear Ni(II)Cu(II) complex with an unusual supramolecular structure: crystal structure and magnetic properties.

Four oxamato-bridged heterotrinuclear Ni(II)Cu(II)Ni(II) complexes of formula ([Ni(bispictn)](2)Cu(pba))(ClO(4))(2).2.5H(2)O (1), ([Ni(bispictn)](2)Cu(pbaOH))(ClO(4))(2).H(2)O (2), ([Ni(cth)](2)Cu(pba))(ClO(4))(2) (3), and ([Ni(cth)](2)Cu(opba))(ClO(4))(2).H(2)O (4) and a binuclear Ni(II)Cu(II) complex of formula [Cu(opba)Ni(cth)].CH(3)OH (5) have been synthesized and characterized by means of elemental analysis, IR, ESR, and electronic spectra, where pba = 1,3-propylenebis(oxamato), pbaOH = 2-hydroxyl-1,3-propylenebis(oxamato), opba = o-phenylenebis(oxamato), bispictn = N,N'-bis(2-pyridylmethyl)-1,3-propanediamine, and cth = rac-5,7,7,12,14,14-hexamethyl-1,4,8,11-tetraazacyclotetradecane. The crystal structures of 1, 3, and 5 have been determined. The structures of complexes 1 and 3 consist of trinuclear cations and perchlorate anions, and that of 5 consists of neutral binuclear molecules which are connected by hydrogen bonds and pi-pi interactions to produce a unique supramolecular "double" sheet. In the three complexes, the copper atom in a square-planar or axially elongated octahedral environment and the nickel atom in a distorted octahedral environment are bridged by the oxamato groups, with Cu.Ni separations between 5.29 and 5.33 A. The magnetic properties of all five complexes have been investigated. The chi(M)T versus T plots for 1-4 exhibit the minimum characteristic of antiferromagnetically coupled NiCuNi species with an irregular spin state structure and a spin-quartet ground state. The chi(M)T versus T plot for 5 is typical of an antiferromagnetically coupled NiCu pair with a spin-doublet ground state. The Ni(II)-Cu(II) isotropic interaction parameters for the five complexes were evaluated and are between 102 and 108 cm(-)(1) (H = -JS(Cu).S(Ni)).

New alternating ferro- and antiferromagnetic one-dimensional complexes. Synthesis, characterization, crystal structure, and magnetic properties of.

The preparation, X-ray crystal structure, and magnetic properties of alternating 1,1- and 1,3-azido-bridged copper(II) complex [Cu(4,4'-dmbpy)(N3)2]n (1, 4,4'-dmbpy = 4,4'-dimethylbipyridine) have been reported. It crystallizes in triclinic system, space group P1, a = 7.9903(1) A, b = 9.3545(9) A, c = 10.754(2) A, alpha = 113.485(1) degrees, beta = 101.399(1) degrees, gamma = 101.897(1) degrees, Z = 2. The magnetic properties of 1 have been investigated in the temperature range 1.5-300 K. Alternating antiferromagnetic (-J = 191.0 cm(-1)) interaction through a 1,3-N3- bridge and ferromagnetic (J = 297.1 cm(-1)) interaction through a 1,1-N3- bridge are obtained for 1 by analyzing the magnetic susceptibility data with the Hamiltonian H = -Jsigma(S2iS2i-1--alphaS2iS2i+1). It's derivatives ([Mn(4,4'-dmbpy)(N3)2]n (2), [Ni(4,4'-dmbpy)(N3)2]n (3), and [Fe(4,4'-dmbpy)(N3)2]n (4) and the heterometallic derivatives [NiMn(4,4'-dmbpy)2(N3)4]n (5) and [CuMn(4,4'-dmbpy)2(N3)4]n (6) have also been synthesized and characterized by electronic and IR spectra. The X-ray powder diffraction and the magnetic properties of 6 have also been discussed.

Role of the pituitary in tumor promotion with ethinyl estradiol in rat liver.

Synthetic estrogens act as tumor promoters in rat liver. Because estrogen treatment markedly increases the secretion of pituitary prolactin, also shown to be a tumor promoter in rat liver, the possibility of a pituitary influence in estrogen promotion was investigated in Wistar rats. In diethylnitrosamine (DEN)-initiated hypophysectomized (hx) female rats, 24 weeks of ethinyl estradiol (EE) administration (500 microg/kg/d, intraperitoneally) did not increase the number of hepatocyte nodules and did not induce hepatocellular carcinoma (HCC) in a 2-year study. Very few placental forms of glutathione-S-transferase (GST-P)-positive foci were observed at the end of EE administration. Estrogen receptor (ER) messenger RNA (mRNA) levels in hx females were 20% of the levels in intact females. EE administration (range, 160-210 microg/kg/d, subcutaneous release pellets) to DEN-initiated intact males and females increased the number and size of hepatocyte foci. A significant increase in HCC frequency was observed in EE-treated females compared with females receiving sham-release pellets, and the latency period for HCC induction was decreased by EE in both males and females. Inhibition of prolactin (PRL) secretion by bromocriptine (Brc) (ParlodelLAR, slow intramuscular release vehicles) during EE treatment decreased the number of foci without affecting their size and markedly prolonged the latency period in both sexes. EE treatment also significantly increased the expression of c-myc, and c-jun, enhanced the levels of growth hormone receptor (GHr) mRNA in females and the levels of ER mRNA in males and "feminized" the expression of the GH-regulated genes cytochrome P450 (CYP), 2C11, CYP 2C12, and GHr in male liver. Brc administration decreased the mRNA levels of the female-predominant CYP 2C12 in EE-treated males but otherwise had no effects. In conclusion, a decreased promotive effect of EE was obtained by decreasing the PRL levels, indicating that estrogens exert at least part of their promotion effects indirectly, by increasing the levels of pituitary PRL.

Reported in vivo splice-site mutations in the factor IX gene: severity of splicing defects and a hypothesis for predicting deleterious splice donor mutations.

Small consensus sequences have been defined for RNA splicing, but questions about splicing in humans remain unanswered. Analysis of germline mutations in the factor IX gene offers a highly advantageous system for studying the mutational process in humans. In a sample of 860 families with hemophilia B, 9% of independent mutations are likely to disrupt splicing as their primary mode of action. This includes 26 splicing mutations reported herein. When combined with the factor IX splice mutations reported by others, at least 104 independent mutations have been observed, 80 of which are single base substitutions within the splice donor and splice acceptor consensus sequences. After analysis of these mutations, the following inferences emerge: (1) the susceptibility of a splice donor sequence to deleterious mutation depends on the degree of similarity with the donor consensus sequence, suggesting a simple "5-6 hypothesis" for predicting deleterious vs. neutral mutations; (2) the great majority of mutations that disrupt the splice donor or splice acceptor sequences result in at least a 100-fold decrement in factor IX coagulant activity, indicating that the mutations at these sites generally function as an on/off switch; (3) mutations that create cryptic splice junctions or that shorten but do not interrupt the polypyrimidine tract in the splice acceptor sequence can reduce splicing by a variable amount; and (4) there are thousands of potential donor-acceptor consensus sequence combinations in the 38-kb factor IX gene region apparently not reduced by evolutionary selective pressure, presenting an apparent paradox; i.e., mutations in the donor and acceptor consensus sequences at intron/exon splice junctions can dramatically alter normal splicing, yet, appropriately spaced, good matches to the consensus sequences do not predispose to significant amounts of alternative splicing.

Crystal structures and magnetic properties of new cyano-bridged two-dimensional grid-like bimetallic assemblies [Ni(tn)2]2[Cr(CN)5((NO)]OH*H2O and [NI(tn)2]2[Co(CN)6]NO3*2H2O (tn=1,3-propanediamine).

Two bimetallic assemblies, [Ni(tn)(2)](2)[Cr(CN)(5)(NO)]OH.H(2)O (1) and [Ni(tn)(2)](2)[Co(CN)(6)]NO(3).2H(2)O (2) (tn = 1,3-diaminopropane), have been prepared and structurally and magnetically characterized. Crystal data for 1 (2): space group P1 (P1), a = 8.698(3) (8.937(2)) A, b = 10.001(2) (9.863(1)) A, c = 10.158(2) (10.064(1)) A, alpha = 87.40(2) (86.064(10)) degrees, beta = 65.10(2) (65.489(10)) degrees, gamma = 81.63(2) (81.572(12)) degrees and Z = 1 (1). Both structures consist of two-dimensional grid-like polycations containing Ni-N triple bond C-M linkages (M = Cr or Co) and counteranions (OH, NO(3)). Magnetic studies of 1 showed that the complex displays a metamagnetic behavior originating from intralayer ferromagnetic and interlayer antiferromagnetic interactions. Long-range antiferromagnetic ordering was observed at T(N) = 3.3 K. Complex 2 exhibits intramolecular ferromagnetic interactions through the diamagnetic N triple bond C-Co-N triple bond C bridges, owing to superexchange involving the empty d(sigma) orbital of the diamagnetic Co(III) ion.