Application of adrenocorticotropic hormone receptor determination and ultrastructural observation of tumor cells in the subtyping of adrenocortical neoplasms / 中国医学科学院学报

<p><b>OBJECTIVE</b>To explore the values of adrenocorticotropic hormone receptor (ACTH-R) determination and ultrastructural observation of tumor cells in the subtyping of adrenocortical neoplasms (ANs).</p><p><b>METHODS</b>The expression of ACTH-R in 87 AN tissues were determined with Polymer immunohistochemical staining, with 10 normal adrenal tissues as the controls. The ultrastructure of the tumor cells was observed using electron microscopy.</p><p><b>RESULTS</b>The positive expression rate of ACTH-R was (80.1 +/- 8.2)%, (53.2 +/- 10.3)%, (63.2 +/- 10.1)%, (83.3 +/- 6.5)%, and (70.1 +/- 7.3)% in the sub-CPA group, CPA group, APA group, NFA group, and NC group, respectively. ACTH-R expression was significantly higher in NFA and sub-CPA groups than in NC group (P = 0.001, P = 0.000), APA group (P = 0.000, P = 0.000), and CPA group (P = 0.000, P = 0.000), and was also significantly different between NC group and APA group (P = 0.039) and between APA group and CPA group (P = 0.037). However, no significant difference was found between NFA group and sub-CPA group (P = 0.325). As shown by the electron microscopy, ANs had some partially similar microscopic features, while different AN subtypes showed differences in the type and amount of secretory granules.</p><p><b>CONCLUSION</b>ACTH-R determination and ultrastructural observation of tumor cells may be helpful for subtyping ANs.</p>

Associations of GR and ACTHR gene polymorphisms with quantitative trait of strain / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To explore association between GR and ACTHR gene polymorphisms and quantitative trait of stress in Chinese Han population.</p><p><b>METHODS</b>Four polymorphic markers (GRA5556G, A5556G, GAGG4534/4536AAAG, promoter T-2C) in GR gene and ACTHR gene were genotyped with PCR-RLFP in 200 healthy Hans. ISTA6.0 and life event stressor questionnaire was used to assess stressors. JSS, SCL-90 and GWB questionnaires were used to quantify the phenotypes of stress. Blood cortical and ACTH levels, and nervous behavior function were measured to assess Physiological strain. CWAI questionnaire was used to assess work ability. Then strain was assessed with Structural equation modeling (SEM).</p><p><b>RESULTS</b>The subjects with GR A5556G genotype (G/A) showed significantly higher plasma cortisol levels, higher psychological stress scores, lower work ability scores and lower plasma ACTH levels compared with the subjects with wild-type (P < 0.01). Psychological stress scores and plasma cortisol levels in the subjects with GR GAGG4534/4536AAAG AG genotype were significantly higher than those in the subjects with wild-type, but the reaction and action sensitivity in the subjects with GR GAGG4534/4536AAAG AG genotype were significantly lower than those in ones with wild-type (P < 0.01). The ACTH level in the subjects with ACTHR promoter T-2C T/T genotype was significantly lower than that in ones with C/C and C/T genotype (P < 0.01). Interaction of GRA5556G and GG4534/4536AAAG with plasma cortisol was positively associated (βs = 0.543, P < 0.01), but with SCL-90 score was negatively associated (βs = -0.374, P < 0.01). Interaction of GRA5556G and GGC6294G with plasma cortisol was correlated (βs = 0.465, P < 0.05). While GR and ACTHR gene variants are the risk factors for psychological strain, physiological strain and decreased work ability (βs are 0.62, 0.43, -0.74, respectively (P < 0.01). While scarce social support, job stressors, negative life stressors and dangerous individual characters are the risk factors for occupational strain, psychological strain, physiological strain and decreased work ability (P < 0.01).</p><p><b>CONCLUSION</b>GRA5556G, GRA5556G, GAGG4534/4536AAAG and ACTHR promoter T-2C variants might be associated with quantitative trait of strain, and GR and ACTHR gene variants with stressors increased the risk for developing strain.</p>

Expressão de genes relacionados à função adrenocortical no estado de caquexia neoplásica / Expression of genes related to the adrenocortical function in the neoplastic cachexia process

A glândula adrenal tem papel fundamental na resposta neuroendócrina, especialmente em situações em que há comprometimento da homeostasia. No processo de caquexia neoplásica, há prejuízo da homeostasia por alterações nutricionais e metabólicas do câncer em estágio avançado, envolvendo a resposta do eixo hipotálamo-hipófise-adrenal. Neste trabalho, foi utilizado um modelo animal de caquexia induzida pelo tumor de Walker-256 em ratos Wistar. Os animais (n=4) foram sacrificados dez dias após a inoculação de células tumorais e a glândula adrenal foi removida. O RNA foi extraído para o estudo da expressão de genes relacionados ao controle da esteroidogênese por RT-PCR semiquantitativa. A análise dos dados demonstrou expressão significativamente reduzida dos genes MC2R (receptor tipo 2 para melacortina), 3ßHSD I (3β-hidroxiesteroidedesidrogenas e tipo I) e TSPO (proteína translocadora) em animais com caquexia neoplásica(valores de P=0,037; 0,0097 e 0,052, respectivamente), revelando falência do córtex da adrenal.

The adrenal gland plays a crucial role in the neuroendocrine response, especially in situations where homeostasis is disturbed. In the neoplastic cachexiaprocess, there is homeostasis impairment by nutritional and metabolic alterations of advanced-stage cancer, involving hypothalamus-pituitary-adrenal axis response. In this assignment, an experimental model of cachexia induced by Walker-256 tumor was performed in Wistar rats. Animals (n=4) were sacrificed 10 days after inoculation of tumor cells, and the adrenal glands were excised. The RNA was isolated for the study of geneexpression related to the steroidogenesis control by semi-quantitative RT-PCR. Dataanalysis showed a significant reduced expression of MC2R (melancortin type 2 receptor), 3ßHSD I (3-beta-hydroxysteroid dehydrogenase type I) and TSPO (translocator protein)genes in animals with neoplastic cachexia (P=0.037, 0.0097 and 0.052, respectively), revealing adrenal cortex failure.

Detection of binding activity and biologic effect of a novel alpha-melanocyte-stimulating hormone analogue / 药学学报

Binding activity and biologic effect of a novel alpha-melanocyte-stimulating hormone analogue were tested on cells transiently expressing the human melanocortin-1 (MC1), MC3, MC4, and MC5 receptors. The human MC1 and MC5 receptor genes were cloned into the expression vector pcDNA3. 1/ myc-his(-) B. The vectors were transferred to HEK-293 cells by the calcium phosphate method. Stable receptor populations were generated using G418 selection (900 microg x mL(-1)) for subsequent bioassay analysis. K(i) values of the novel alpha-MSH analogue for MC1, MC3, MC4, and MC5 receptors were obtained in competition with [125I]-NDP-MSH for binding studies. The cyclic AMP level was tested by using [3H]-cyclic AMP kit. It is showed that K(i) values of the novel alpha-MSH analogue for MC1, MC3, MC4, and MC5 receptors were (0.159 +/- 0.040), (35.430 +/- 6.743), (19.293 +/- 2.780) and (2.230 +/- 0.670) nmol L(-1), respectively. Its EC50 values for MC1, MC3, MC4, and MC5 receptors were (0.45 +/- 0.07), (7.80 +/- 0.65), (2.55 +/- 0.23) and (0.33 +/- 0.09) nmol L(-1), respectively. In these tests, the novel alpha-MSH analogue is a MC1R and MC5R selective agonist.

Controle neuroendócrino do peso corporal: implicaçöes na gênese da obesidade / Neuroendocrine control of food intake: implications in the genesis of obesity

O peso corporal é regulado por uma interaçäo complexa entre hormônios e neuropeptídeos, sob o controle principal de núcleos hipotalâmicos. Mutaçöes nos genes de hormônios e neuropeptídeos, de seus receptores ou de elementos regulatórios, têm sido descritas na espécie humana, mas säo tidas como raras, näo explicando as formas mais comuns de obesidade. No entanto, o estudo destas mutaçöes tem propiciado um grande avanço nos conhecimentos sobre a base genética e a fisiopatologia da obesidade, possibilitando o estudo e abrindo perspectivas para o desenvolvimento de novas modalidades terapêuticas. Recentemente, demonstrou-se que mutaçöes no receptor 4 da melanocortina podiam ser encontradas em até 5 por cento dos casos de obesidade severa, representando até o presente momento a forma mais prevalente de obesidade monogênica na espécie humana. Nesta revisäo, säo discutidas as diversas mutaçöes descritas nos seres humanos de elementos da rede neuroendócrina de controle do peso corporal, bem como as implicaçöes dos mesmos na gênese da obesidade

The Effect of alpha MSH Analogues on Rat Bones

Melanocortin is the downstream mediator of leptin signaling and absence of leptin signaling in ob/ob and db/db mice revealed the enhancement of bone formation through the central regulation. While alpha-melanocyte-stimulating hormone (alpha MSH) inhibits the secretion of interleukin-1 alpha and tumor necrosis factor-alpha from the inflammatory cells, alpha MSH can also enhance clonal expansion of pro B cells linked to stimulation of osteoclastogenesis. Therefore, we tested the effect of melanocortin on bones. alpha MSH analogues [6His] alpha MSH-ND and [6Asn] alpha MSH-ND were synthesized and the radio-ligand receptor binding- and cyclic AMP generating activity were analyzed in China Hamster Ovary cell line over- expressing melanocortin receptors. The EC50 of [6His] alpha MSH-ND measured from melanocortin-1, 3, 4 and 5 receptors were 0.008 0.0045, 1.523 0.707, 0.780 0.405, and 250.320 42.234 nM, respectively, and the EC50 of [6Asn] alpha MSH-ND were 16.8 6.94, 271.8 21.95, 8.0 1.21, and 1132.5 635.46 nM, respectively. Four weeks after the subcutaneous injection of the analogues, the body weights in the [6His] alpha MSH-ND and the [6Asn] alpha MSH-ND treated groups (346.0 20.63 g vs. 350.0 13.57 g) were lower than that of the vehicle treated group (375.8 17.31 g, p 0.05). There was no difference in the total femoral BMD measured by dual x-ray absorptiometry among the three groups. Among the three groups, there were no differences in the total numbers of crystal violet positive- or alkaline phosphatase positive colonies, in the expression of Receptor Activator of Nuclear Factor Kappa-B ligand on the tibia and the total number of multinucleated osteoclast-like cells differentiated from primary cultured bone marrow cells. From the above results, no evidence of bone gain or loss was found after treatment of the alpha MSH analogues peripherally.

Proliferative signaling initiated in ACTH receptors

This article reviews recent results of studies aiming to elucidate modes of integrating signals initiated in ACTH receptors and FGF2 receptors, within the network system of signal transduction found in Y1 adrenocortical cells. These modes of signal integration should be central to the mechanisms underlying the regulation of the G0->G1->S transition in the adrenal cell cycle. FGF2 elicits a strong mitogenic response in G0/G1-arrested Y1 adrenocortical cells, that includes a) rapid and transient activation of extracellular signal-regulated kinases-mitogen-activated protein kinases (ERK-MAPK) (2 to 10 min), b) transcription activation of c-fos, c-jun and c-myc genes (10 to 30 min), c) induction of c-Fos and c-Myc proteins by 1 h and cyclin D1 protein by 5 h, and d) onset of DNA synthesis stimulation within 8 h. ACTH, itself a weak mitogen, interacts with FGF2 in a complex manner, blocking the FGF2 mitogenic response during the early and middle G1 phase, keeping ERK-MAPK activation and c-Fos and cyclin D1 induction at maximal levels, but post-transcriptionally inhibiting c-Myc expression. c-Fos and c-Jun proteins are mediators in both the strong and the weak mitogenic responses respectively triggered by FGF2 and ACTH. Induction of c-Fos and stimulation of DNA synthesis by ACTH are independent of PKA and are inhibited by the PKC inhibitor GF109203X. In addition, ACTH is a poor activator of ERK-MAPK, but c-Fos induction and DNA synthesis stimulation by ACTH are strongly inhibited by the inhibitor of MEK1 PD98059.

Role of ACTH receptor in adrenocortical tumor formation

Adrenocorticotrophin (ACTH) is the major regulatory hormone of steroid synthesis and secretion by adrenocortical cells. The actions of ACTH are mediated by its specific membrane receptor (ACTH-R). The human ACTH-R gene was recently cloned, allowing systematic determination of its sequence, expression and function in adrenal tumorigenesis. The presence of oncogenic mutations of the ACTH-R gene in adrenocortical tumors has been reported. Direct sequencing of the entire coding region of the ACTH-R gene of sporadic adrenocortical adenomas and carcinomas did not reveal constitutive activating mutations, indicating that this mechanism is not frequent in human adrenocortical tumorigenesis. Recent studies demonstrated allelic loss of the ACTH-R gene in a subset of sporadic adrenocortical tumors using a PstI polymorphism located in the promoter region of the ACTH-R gene. Loss of heterozygosity of the ACTH-R was analyzed in 20 informative patients with a variety of benign and malignant adrenocortical tumors. Three of them showed loss of heterozygosity of the ACTH-R gene. In addition, Northern blot experiments demonstrated reduced expression of ACTH-R mRNA in these three tumors with loss of heterozygosity, suggesting the functional significance of this finding at the transcriptional level. Deletion of the ACTH-R gene seems to be involved in a subset of human adrenocortical tumors, contributing to cellular dedifferentiation.

The expression of the ACTH receptor

Adrenal glucocorticoid secretion is regulated by adrenocorticotropic hormone (ACTH) acting through a specific cell membrane receptor (ACTH-R). The ACTH-R is a member of the G protein superfamily-coupled receptors and belongs to the subfamily of melanocortin receptors. The ACTH-R is mainly expressed in the adrenocortical cells showing a restricted tissue specificity, although ACTH is recognized by the other four melanocortin receptors. The cloning of the ACTH-R was followed by the study of this gene in human diseases such as familial glucocorticoid deficiency (FGD) and adrenocortical tumors. FGD is a rare autosomal recessive disease characterized by glucocorticoid deficiency, elevated plasma ACTH levels and preserved renin/aldosterone secretion. This disorder has been ascribed to an impaired adrenal responsiveness to ACTH due to a defective ACTH-R, a defect in intracellular signal transduction or an abnormality in adrenal cortical development. Mutations of the ACTH-R have been described in patients with FGD in segregation with the disease. The functional characterization of these mutations has been prevented by difficulties in expressing human ACTH-R in cells that lack endogenous melanocortin receptor activity. To overcome these difficulties we used Y6 cells, a mutant variant of the Y1 cell line, which possesses a non-expressed ACTH-R gene allowing the functional study without any background activity. Our results demonstrated that the several mutations of the ACTH-R found in FGD result in an impaired cAMP response or loss of sensitivity to ACTH stimulation. An ACTH-binding study showed an impairment of ligand binding with loss of the high affinity site in most of the mutations studied.

Rol del tejido adiposo / Adipose tissue role

Currently the research on adipose tissue has yielded same insights of itïs function. On the embrionary state the differentiation between white adipose tissue and brown adipose tissue and brown adipose tissue signals different functions. The presence of multiple receptors and actions define itïs autocrine, paracline and endocrine roles

Induction of fos and jun proto-oncogenes by the adrenocorticotropic hormone (ACTH): possible biological implications

ACTH induces the expression of fos and jun proto-oncogene family members in the mouse Y-1 adrenocortical cell fine. PMA (phorbol-12-myristate-l3-acetate) closely mimics these inductive effects of ACTH. On the other hand, cAMP derivatives are not effective in inducing the fos and jun genes. These results suggest that ACTH receptors are likely to activate signaling routes other than the classical cAMP/protein kinase A in order to induce FOS and JUN proteins. We hypothesize that induction of FOS and JUN proteins is likely to be important in the trophic response of adrenocortical cells to ACTH.