The human forebrain has discrete estrogen receptor alpha messenger RNA expression: high levels in the amygdaloid complex

Estrogen is considered to play an important role in neuropsychiatric disorders and the estrogen receptors mediate the action of the hormone. In the present study, the messenger RNA expression pattern of the estrogen receptor alpha subtype was identified in the post mortem human brain. High stringent in situ hybridization histochemistry was performed using a riboprobe specific for the estrogen receptor alpha subtype. The human brain was mainly characterized by abundant estrogen receptor alpha messenger RNA expression in the amygdala and hypothalamus, but labeling (lower) was also found in the extended sublenticular amygdala, cerebral cortex, and hippocampus. In the amygdala, the estrogen receptor alpha messenger RNA was preferentially expressed in medially-localized nuclei suggesting that estrogen regulates distinct human amygdala-mediated functions. The Cynomologous monkey brain was also examined in the present study and a similar distribution of the estrogen receptor alpha messenger RNA signal was observed in the human and monkey brain. However, the primate expression pattern differed in part from the known distribution in the rat. The current results show that estrogen receptor alpha messenger RNA is expressed in discrete areas of the human brain not only related to neuroendocrine function, but also emotion, memory, and cognition, which is consistent with the hypothesized involvement of estrogen in schizophrenia, affective disorders, and Alzheimers disease.     

Cyclin D1 amplification and expression in human breast carcinoma: correlation with histological prognostic markers and oestrogen receptor expression

Aims—To study the amplification of the Cyclin D1 gene (CCND1) in human breast carcinoma; to relate this to Cyclin D1 protein expression; to relate these parameters to recognised pathological prognostic factors, including oestrogen receptor (ER) status.     

Hepcidin messenger RNA expression in human lymphocytes

Hepcidin regulates intracellular iron levels by interacting with and promoting the degradation of ferroportin, a membrane protein and the only known cellular iron exporter. Studies of hepcidin expression and regulation have focused on its effects in innate immunity and as a regulator of systemic iron metabolism. In the present study we characterized the expression of hepcidin messenger RNA (mRNA) in human peripheral blood mononuclear cells (PBMCs) with a focus on peripheral blood lymphocytes (PBLs). We found that (1) all human PBMCs analyzed express basal hepcidin mRNA levels; (2) hepcidin mRNA expression increases after T‐lymphocyte activation; (3) expression by PBLs increases in response to challenge by holotransferrin (Fe‐TF) and by ferric citrate in vitro; (4) the Fe‐TF‐mediated up‐regulation of hepcidin decreases ferroportin expression at the cytoplasmic membrane of PBLs; and (5) silencing of tumour necrosis factor‐α (TNF‐α) abrogates the effect of Fe‐TF. In summary, we show that hepcidin expression determines intracellular iron levels by regulating the expression of ferroportin, as described in other cells, and that inappropriately low expression of hepcidin impairs normal lymphocyte proliferation. The results establish hepcidin as a new player in lymphocyte biology.     

Mean nuclear area and metallothionein expression in ductal breast tumors: correlation with estrogen receptor status.

Breast carcinoma is the most common malignancy in women. Estrogen is an important growth factor for breast tumor that plays an important role in regulating the proliferation and differentiation of normal and malignant mammary epithelial cells. Nuclear morphometry and metallothioneins (MTs) are indicators of proliferation that have been used as predictors of prognosis in many tumors. The present study aimed to study mean nuclear area (MNA) and MT; estrogen receptor (ER) expression in fibroadenoma (FA), ductal carcinoma in situ (DCIS), and infiltrating ductal carcinoma (IDC) of the breast. Also MNA and MT expression will be correlated with histologic grade and ER status in breast carcinoma. Breast tissues from 18 patients with FA, 10 patients with DCIS, and 40 patients with IDC were used in this study. MNA and MT expression; as proliferation markers; were investigated and correlated with ER status. All cases of FA, 7 out of 10 cases (70%) of DCIS and 23 out of 40 cases (57.5%) of IDC were positive for ER. MNA of cancer cells was significantly larger than that of normal and benign breast tissue. A significant direct correlation was found between MNA and histologic grades. MNA of ER-negative carcinomas was significantly larger than that of ER-positive tumors. In normal and benign breast tissue, myoepithelial cells consistently expressed the MT protein. Four out of 10 DCIS cases (40%) and 24 out of 40 cases of IDC cases (60%) were positively stained for MT. MT positivity was directly correlated with histologic grade of IDC. There was a highly significant inverse correlation between MT and ER overexpression. From this study, it is concluded that in invasive ductal carcinoma of the breast, the large MNA and MT overexpression are correlated with histologic grades and ER negativity. Therefore, large MNA and MT overexpression may be possible important indicators for more aggressive and less differentiated breast carcinoma.     

正常及去势大鼠骨组织内雌激素受体mRNA表达水平的研究

通过切除大鼠卵巢复制实验性骨质疏松动物模型，从小块胫骨组织内提取ＲＮＡ，对正常对照、切除卵巢及雌激素治疗组的标本进行逆转录。聚合酶链反应（ＲＴ－ＰＣＲ）检测，直接从生理状态下的成熟骨组织中检测出雌激素受体（ＥＲ）ｍＲＮＡ的表达，并经测序证实。实验结果还显示切除卵巢大鼠的ＥＲ表达水平明显降低，而予以雌激素治疗组的ＥＲ表达接近正常。提示ＥＲ的表达依赖于雌激素的存在，而雌激素通过ＥＲ在骨代谢中起作用。     

Expression of somatostatin type 2A receptor correlates with estrogen receptor in human breast carcinoma

Somatostatin type 2A receptor (sstr2A) has been shown to be directly involved in the transduction of antiproliferative effects and also to be the most predominant sstr subtype in human normal breast epithelium, as well as in human breast carcinoma. We investigated the immunoreactivity of sstr2A in 34 cases of human breast carcinoma and correlated these findings with the immunoreactivity of the estrogen receptor (ER), epidermal growth factor receptor (EGFR), transforming growth factor α(TGFα) and insulin like growth factor I (IGF-I). We detected sstr2A immunoreactivity in normal mammary tissue, and in 27 of 34 (79%) breast carcinomas. The sstr2A immunoreactivity was localized on the cellular membrane, however, weak cytoplasmic immunoreactivity was also observed. Sstr2A immunoreactivity was heterogenously distributed in the whole tumor section. There was a statistically significant correlation between sstr2A and ER immunoreactivity in the same tumor. No statistically significant correlation was found between sstr2A immunoreactivity and immunoreactivity for EGFR, TGFα and IGF-I or the patients' age.     

Keratin expression in the normal breast and in breast carcinoma

The immunohistochemical reactivities of 69 cases of breast carcinoma were examined on methacarn-fixed, paraffin-embedded sections using eight different monoclonal antibodies which recognize one or a few keratin polypeptides. In the normal breast, the monoclonal antibodies RPN1162, RPN1165 and AE1 stained almost all the luminal cells but not the basal (myoepithelial) cells. The monoclonal antibodies 35BH11, M20, CK5 and CK8.12 stained only a subset of the luminal cells. In contrast, 312C8-1 stained basal cells but not luminal cells. All the tumour specimens reacted with AE1, while over 80% of them also reacted with 35BH11 (57/69), CK5 (57/69) and RPN1165 (55/69); 30% reacted with CK8.12 (21/69) and 16% with RPN1162 (11/69). Basal cell-specific keratin, as defined by 312C8-1, was detected in only 1% of cases (1/69). Monoclonal antibodies to different keratin polypeptides may be of use in the characterization and subdivision of breast cancer.     

Clinicopathological value of somatostatin type 2A and estrogen receptor immunoreactivity in human breast carcinoma

Somatostatin type 2A (sstr2A) and estrogen receptor (ER) are interrelated regulatory receptors present in normal breast epithelium and in a population of breast carcinomas. ER mediates growth stimulatory effects of estrogens whereas sstr2A mediates growth inhibitory actions of somatostatin. However, much work has been devoted to elucidate the biological role of ER, little is known about sstr2A in breast cancer. In the present study we examined immunoreactivity of sstr2A and ER in 64 breast carcinomas in correlation with tumor size and histological grade (HG), presence of lymph node metastasis (LNM), Nottingham prognostic index (NPI), and the patients' age. ER and sstr2A immunoreactivity were present in 78% and 63% of the breast carcinomas, respectively. Ninety percent of tumors immunoreactive for sstr2A were simultaneously immunoreactive for ER. ER immunoreactivity correlated significantly with lower HG (p=0.03) and better NPI (p=0.02). sstr2 A immunoreactivity correlated significantly with lower HG (p=0.012) but not with NPI (p=0.26). There was no correlation of sstr2A immunoreactivity and tumor size, patients' chronological age or LNM. The results confirm prognostic value of ER immunohistochemistry in breast carcinoma. However sstr2A cannot substitute ER for prognostic evaluation, sstr2A immunoreactivity being significantly associated with lower HG seems to represent an independent prognostic factor in breast carcinoma.     

Immunohistochemical expression of gonadotropin releasing hormone receptor in human breast carcinoma

Gonadotropin releasing hormone (GnRH) analogs can cause regression of hormone-dependent breast carcinomas via the specific GnRH receptor (GnRH-R). In an attempt to obtain a better understanding of GnRH actions in human breast carcinoma, the expression of GnRH-R was examined immunohistochemically in 58 invasive ductal carcinomas and correlated with various clinicopathological parameters. GnRH-R was immunolocalized in the cytoplasm of carcinoma cells in 37 of 58 invasive ductal carcinoma cases (64%). Immunoreactivity for GnRH-R was also detected focally in the cytoplasm of morphologically normal glandular epithelia adjacent to the carcinoma. A significant correlation was observed between the immunohistochemical expression of GnRH-R and estrogen receptor labeling index (LI; P = 0.030) or progesterone receptor LI (P = 0.0074). There was a significant inverse correlation between GnRH-R immunoreactivity and Ki-67 LI (P = 0.012). No significant correlations were detected between GnRH-R and other clinicopathological parameters, including patient age, menopausal status, stage, tumor size, lymph node status, histological grade and prognosis. This study indicates that GnRH-R is widely distributed in human breast carcinoma cells and regulates GnRH actions locally. Breast carcinomas positive for GnRH-R maintain some hormonal regulatory mechanisms, and GnRH actions may lead to a low proliferative rate in human breast carcinoma.     

Immunohistochemical analysis of estrogen receptor alpha, estrogen receptor beta and progesterone receptor in normal human endometrium

The endometrium expresses estrogen (ER) and progesterone receptors (PR), which are involved in autocrine and paracrine regulation processes in response to estrogen and progesterone. The aim of the present study was to evaluate immunohistochemical distribution patterns of estrogen receptor alpha (ER alpha), estrogen receptor beta (ER beta) and PR in normal human endometrial tissue with the use of monoclonal antibodies. Human endometria were obtained from 17 premenopausal patients undergoing surgery for non-malignant diseases and were classified to be in proliferative, early secretory and late secretory phases by histological and anamnestical means. Distribution patterns of the steroid receptors were evaluated using the IRS-score and the Mann-Whitney rank-sum test was used to compare the means. Correlation was assessed with the Spearman factor and linear regression analysis. ER alpha and PR expression decreased significantly (p<0.05) in glandular epithelium from the proliferative to the late secretory phase. ER beta expression showed a similar significant decrease (p<0.05), although staining intensity was lower than that of ER alpha. A significant correlation between expression of all three steroid receptors was observed (p<0.001). Distribution patterns of ER alpha, ER beta and PR in normal human endometrium showed a cyclic variation during the menstrual cycle. A significant correlation between expression of ER alpha, ER beta and PR was also demonstrated using regression analysis, indicating dependence of expression of these three steroid receptors. The present study shows the presence of steroid receptors in human endometrial epithelium, indicating that these cells respond to estrogen and progesterone and thus playing a significant role in endometrial physiology.     

The expression of nuclear estrogen receptor and its relation to cytoplasmic estrogen receptor in breast cancer]

Sixty specimens of breast cancer were assayed with ER-monoclonal antibody by immunocytochemical staining. Twenty-nine (48.33%) were nuclear estrogen receptor positive (ERn+). The number of ERn+ cancer cells decreased in the following sequence: mucinous carcinoma, invasive lobular carcinoma, invasive ductal carcinoma, and papillary adenocarcinoma. Two apocrine carcinomas were ERn-. ERn+ rate was higher in patients over 60 years of age (P less than 0.05). The amount of ERn+ cells was much greater in cancer cells than in the surrounding benign epithelial cells. This phenomenon may indicate that malignant cells are more hormone-dependent than benign cells. The results of immunocytochemical staining and steroid binding assay were compared. By immunocytochemical staining, twenty-four of fifty-seven cases (42.11%) were ERn+ and ERc+. Sixteen cases (28.07%) were ERn- and ERc-. This study showed that in a number of breast cancers ERs were positive in both cytoplasms and nuclei and the concordant rate was 70.18%. In the remaining cases 13 (22.81%), ERs were positive in the cytoplasm, and in 4 cases (7.02%) positive in nuclei only. Additionally, fifty-two out of 60 cases were assayed by 3H-estradiol and 3H-R5020 by means of steroid binding assay. Twenty-seven cases of them (51.92%) showed ERc+ and PRc+, and seventeen cases (32.69%) were ERc- and PRc-. Their correspondent rate was 84.61%.     

Oestrogen receptor expression in the normal and pre-cancerous breast

As oestrogen is associated with most of the epidemiological risk factors for breast cancer, the number and distribution of oestrogen receptor positive (ER+) cells could have a bearing on the development of the disease. ER+ cells were thus studied in the normal breast and in the spectrum of in situ proliferations which range from non-atypical hyperplasia to in situ carcinoma and are associated with different levels of risk for developing breast cancer. In the normal pre-menopausal breast, ER+ cells comprised the minority and were distributed singly, being surrounded by oestrogen receptor negative (ER−) cells. ER+ cells showed a statistically significant increase with age, reaching a plateau after the menopause, and the increase was associated with a tendency for positive cells to become contiguous in patches of variable size. A small proportion of lobules showing involutional change comprised over 90 per cent ER+ cells. The significance of this feature is not clear but no evidence was found that it was pre-cancerous. The percentage of ER+ cells was slightly increased in hyperplasia of usual type (non-atypical hyperplasia, HUT) and the relationship to age was maintained. The staining pattern was variable; in some lesions ER+ cells were surrounded by ER− cells whereas in others there were contiguous groups of positive cells sometimes accounting for more than 90 per cent of cells in the lesion. In contrast, all cases of atypical ductal hyperplasia (ADH), lobular in situ neoplasia (LIN) and ductal carcinoma in situ (DCIS) exhibited positivity of contiguous cells accounting for the majority in the lesions. Furthermore, the relationship between ER+ cell numbers and age was lost in these lesions, indicating autonomy of ER expression or of proliferation of cells expressing the receptor. It is hypothesized that this dysregulation of receptor expression or of ER+ cell numbers at the ADH stage may be the precursor of abnormal expression of cyclins and other cell cycle control proteins which have been shown first to appear in DCIS. Copyright © 1999 John Wiley & Sons, Ltd.     

Melanocortin-4 receptor messenger RNA expression is up-regulated in the non-damaged striatum following unilateral hypoxic-ischaemic brain injury

Melanocortin peptides (alpha-melanocyte-stimulating hormone, adrenocorticotropin and fragments thereof) have been shown to have numerous effects on the central nervous system, including recovery from nerve injury and retention of learned behaviour, but the mechanism of action of these peptides is unknown. A family of five melanocortin receptors have recently been discovered, two of which (melanocortin-3 and melanocortin-4 receptors) have been mapped in the rat brain. We have tested the hypothesis that the expression of one or more of the messenger RNAs for three melanocortin receptors (melanocortin-3, melanocortin-4 and melanocortin-5 receptors) would be altered in rat brain following unilateral transient hypoxic-ischaemic brain injury. In this study, using in situ hybridization, we show that melanocortin-4 receptor messenger RNA was up-regulated in the striatum in the non-damaged hemisphere within 24 h after severe hypoxic-ischaemic injury compared with control brains (P<0.05). In a small group of animals, this induction was not blocked by treatment with the anticonvulsant, carbamazepine. Expression of melanocortin-3 receptor messenger RNA in the brain was not altered in this hypoxic-ischaemic injury model and melanocortin-5 receptor messenger RNA was not detected in either control or hypoxic-ischaemic injured rat brains. We hypothesize that the up-regulation of melanocortin-4 receptor messenger RNA expression in the contralateral striatum may be involved in transfer of function to the uninjured hemisphere following unilateral brain injury.     

Differential display and cloning of messenger RNA from human normal nasal epithelial cells versus nasopharyngeal carcinoma cell lines

Carcinogenesis is a multi-process event that has been characterized both by activation of cellular oncogenes and by loss of function of tumor suppressor genes. However, no systemic study has been performed to understand the involvement of oncogenes and tumor suppressor genes in the oncogenesis of nasopharyngeal carcinoma (NPC). Differential display was performed to identify genes specifically expressed in normal nasal epithelial cells or NPC cell line HONE-1. Using Ltk3 and T11CA as primers, a 379-bp cDNA fragment (CN3) obtained from normal nasal epithelial cells was able to show specificity by northern blot analysis. A 3.5-kb mRNA was detected in normal nasal epithelial cells but not in NPC cell line HONE-1 by using 32P-end-labeled CN3 fragment as a probe. Sequence analysis of the 379 bp cDNA fragment indicated unique sequences from nts 1 to 230. Nts 231 to 379 are Alu-like sequences. Northern blot analysis using 32p-labeled PCR product amplified from nts 36 to 222 of CN3 cDNA fragment was also able to detect the 3.5-kb mRNA in normal nasal epithelial cells but not in HONE-1 and two other NPC cell lines NPC-TWO1, NPC-TWO4.     

The expression of parathyroid hormone messenger RNA in normal and abnormal parathyroid tissue.

The distribution and expression of preproparathyroid hormone (PTH) mRNA were investigated in parathyroid tissue from 57 parathyroidectomy specimens. PTH mRNA was detected by in situ hybridization using digoxigenin-labelled oligonucleotide probes. Cell morphology was seen to correlate with PTH mRNA expression. Strong expression of PTH mRNA was confined to cells which on haematoxylin and eosin staining had large vesicular nuclei. These included both vacuolated and non-vacuolated cells. Chief cells with small dark nuclei and scanty cytoplasm had little or no expression. In both adenoma and chief cell hyperplasia, the striking difference from normal was the greatly increased proportion of cells expressing PTH mRNA. In adenomas, the rim of uninvolved parathyroid tissue showed PTH mRNA expression similar to that of normal parathyroid. In hyperplasia, there was frequently concordance of staining within individual nodules. The findings establish morphological criteria for activity of parathyroid tissue and support current concepts of the different pathogenesis of hyperplasia and adenoma. The expression of PTH mRNA in oxyphil change and parathyroid carcinoma was also investigated.     

Prolactin receptor expression in gynaecomastia and male breast carcinoma

Aims:  Despite the well-established function of prolactin (PRL) in normal breast development, its role in breast cancer pathogenesis is still controversial. PRL activity is dependent on the activation of a transmembrane protein, the PRL receptor (PRLR). The aim was to evaluate and compare PRLR expression in gynaecomastia and male breast carcinoma (MBC).
Methods and results:  PRLR expression was detected immunohistochemically in 30 cases of gynaecomastia and 30 cases of MBC. The whole series was also assessed for oestrogen receptors (ER), progesterone receptors (PR) and androgen receptors (AR). A cut-off of 10% was used as the criterion for positivity. Histological type and tumour differentiation were evaluated. Pathological stage was assessed [Tumour Node Metastasis (TNM)-International Union Against Cancer system]. Statistical analysis was performed with Fisher's exact test. PRLR positivity was seen in 20% of gynaecomastia cases and in 60% of MBC cases ( P  = 0.003). In gynaecomastia immunoreactivity was predominantly observed in luminal cell borders, whereas in MBC the reactivity was heterogeneous and mainly cytoplasmic. There was no statistically significant correlation between PRLR expression and ER, PR, AR, pTNM, or histological grade.
Conclusions:  PRLR is significantly more expressed in MBC than in gynaecomastia, and with different patterns of reactivity, suggesting a role for PRL in male breast carcinogenesis.