LI-cadherin gene expression during mouse intestinal development.
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LI-cadherin (Liver-Intestine cadherin) is a member of a subclass (7-D cadherins) within the cadherin superfamily. Although its cellular function as a cell-cell adhesion molecule has been demonstrated in cell culture studies, its physiological function still needs to be explored in the intact organism. After isolating the cDNA for mouse LI-cadherin, we generated specific antibodies against the overexpressed protein and studied its expression pattern in adult mouse tissues and mouse embryos. The mouse LI-cadherin sequence is 91% identical to the sequence of rat LI-cadherin and exhibits the same structural features described for rat LI-cadherin. In mouse adult tissue, LI-cadherin is expressed in the intestine and in small amounts in the spleen. In contrast to rat, Mouse LI-cadherin was not expressed in liver. During mouse embryogenesis, LI-cadherin expression begins at embryonic day 12.5. With the exception of transient expression in the urogenital sinus and the common bile duct on day 13.5, LI-cadherin was found exclusively in the intestinal epithelium. Its expression coincides with the formation of intestinal villi, a developmental stage that includes major tissue remodeling, growth, and differentiation. LI-cadherin is expressed along the entire anterior-posterior axis of the developing intestine as well as along the entire villus axis once villi begin to form. LI-cadherin occupies all cell surfaces of the deeper layers of the epithelium, distributing to basolateral surfaces only in the cells of the outer epithelial layer. LI-cadherin was found to be always co-expressed with E-cadherin. (+info)
A human prostatic epithelial model of hormonal carcinogenesis.
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The effects of stromal and hormonal environment on the immortalized but nontumorigenic human prostatic epithelial cell line BPH-1 were investigated in an in vivo model. BPH-1 cells were recombined with rat urogenital sinus mesenchyme (UGM), and the tissue recombinants were grafted to the renal capsule of adult male athymic mouse hosts. BPH-1 + UGM recombinants formed solid branching epithelial cords with a well-defined basement membrane. The cords canalized to form ductal structures. The mesenchymal cells formed thick sheets of well-differentiated smooth muscle surrounding the epithelium, reinforcing the idea that the epithelium dictates the patterning of prostatic stromal cells. When hosts carrying BPH-1 + UGM tissue recombinants were exposed to testosterone propionate and 17-beta-estradiol (T + E2), the tissue recombinants responded by forming invasive carcinomas, demonstrating mixed, predominantly squamous as well as adenocarcinomatous (small acinar and mucinous) differentiation. When either untreated or T + E2-treated hosts were castrated, epithelial apoptosis was observed in the grafts. When tumors were removed and regrafted to fresh hosts they grew rapidly. Tumors were serially regrafted through six generations. Histologically these tumors consisted largely of focally keratinizing squamous cell carcinoma with high-grade malignant cytological features. BPH-1 cells grown in the absence of UGM survived at the graft site but did not form tumors or organized structures. This behavior was not influenced by the presence or absence of T + E2 stimulation. These data show that an immortalized, nontumorigenic human prostatic epithelial cell line can undergo hormonal carcinogenesis in response to T + E2 stimulation. In addition, the data demonstrate that the stromal environment plays an important role in mediating hormonal carcinogenesis. (+info)
Localization of glucocorticoid receptor interacting protein 1 in murine tissues using two novel polyclonal antibodies.
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OBJECTIVE: Glucocorticoid receptor interacting protein 1 (GRIP1) is a coactivator that binds to the nuclear hormone receptors in a ligand-dependent manner and mediates transcriptional activation of the target genes. The aim of this study was to investigate GRIP1 expression in various murine tissues and whether the protein is nuclear, cytoplasmic, or both. DESIGN: Two novel polyclonal antibodies against amino acids 34-47 and 468-481 of GRIP1 were raised and characterized in order to study the GRIP1 expression with immunohistochemistry. RESULTS: Transient transfection studies with COS cells showed a clearly nuclear staining pattern and also immunohistochemical localization of GRIP1 was mainly nuclear, but cytoplasmic expression was seen as well. GRIP1 was expressed in epithelial cells of the submandibular gland, gastrointestinal tract, pancreas, kidney, uterus, mammary gland, testis, prostate, trachea, lungs and adrenal gland. GRIP1 was also detected in stromal cells of colon, rectum, urinary bladder, vagina, uterus, mammary gland and trachea, and to a lesser extent in esophagus, ureter, urethra, thymus and spleen. Smooth muscle cells of the gastrointestinal and urinary tract, uterus, epididymis, prostrate and bronchioles expressed GRIP1. Blood vessels of many organs, capsule of the kidney and prostate, mesovarium, adipocytes of the mammary gland, pericardium and cartilage of the trachea were also GRIP1-positive. Liver, thyroid gland and striated muscle did not express GRIP1. CONCLUSIONS: GRIP1 was expressed in a wide variety of murine organs, and expression varied between cell types and organs. In addition to mainly nuclear localization of endogenous GRIP1, cytoplasmic expression was seen as well. (+info)
Brown fat UCP1 is specifically expressed in uterine longitudinal smooth muscle cells.
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Until now, uncoupling protein 1 (UCP1) was considered as unique to brown adipocytes. It supports a highly regulated uncoupling of oxidative phosphorylation that is associated with diet as well as with non-shivering thermogenesis. Here we report that UCP1 is not specific to brown adipocytes and can be expressed in longitudinal smooth muscle layers. In the uterus, this conclusion was drawn from different convergent data. A specific antibody against mouse UCP1 revealed, in mitochondrial fractions, a protein with the same molecular weight as brown fat UCP1. Sensitive and specific reverse transcriptase-polymerase chain reaction detected a mRNA whose sequence was totally homologous to that of brown fat UCP1 mRNA. Antibody against UCP1 as well as a UCP1 antisense probe specifically stained uterine longitudinal smooth muscles. UCP1 was also expressed in longitudinal smooth muscle of digestive and male reproductive tracts but was never expressed in other types of smooth muscle, including those of arterial vessels. In uterine tract, UCP1 content was increased after cold exposure or beta-adrenergic agonist treatment. It was also up-regulated during the postovulatory period after sexual cycle synchronization. Its content transiently increased during gestation and decreased markedly after birth. These regulations strongly argue about a role for UCP1 in thermogenesis as well as in relaxation of longitudinal smooth muscle layers. (+info)
Mechanisms of estrogen action.
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Our appreciation of the physiological functions of estrogens and the mechanisms through which estrogens bring about these functions has changed during the past decade. Just as transgenic mice were produced in which estrogen receptors had been inactivated and we thought that we were about to understand the role of estrogen receptors in physiology and pathology, it was found that there was not one but two distinct and functional estrogen receptors, now called ER alpha and ER beta. Transgenic mice in which each of the receptors or both the receptors are inactive have revealed a much broader role for estrogens in the body than was previously thought. This decade also saw the description of a male patient who had no functional ER alpha and whose continued bone growth clearly revealed an important function of estrogen in men. The importance of estrogen in both males and females was also demonstrated in the laboratory in transgenic mice in which the aromatase gene was inactivated. Finally, crystal structures of the estrogen receptors with agonists and antagonists have revealed much about how ligand binding influences receptor conformation and how this conformation influences interaction of the receptor with coactivators or corepressors and hence determines cellular response to ligands. (+info)
Clinical utility of a nested nucleic acid amplification format in comparison to viral culture for the diagnosis of mucosal herpes simplex infection in a genitourinary medicine setting.
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BACKGROUND: Nested nucleic acid amplification tests are often thought too sensitive or prone to generating false positive results for routine use. The current study investigated the specificity and clinical utility of a routine multiplex nested assay for mucosal herpetic infections. METHODS: Ninety patients, categorised into those clinically diagnosed to (a) have and (b) not have herpetic infection, were enrolled. Swabs from oral and ano-genital sites were assayed by the nested assay and culture and the results assessed against clinical evaluation for diagnosing herpetic infections; cell content was also recorded. RESULTS: Twenty-six and 64 patients were thought to (a) have and (b) not have mucosal herpetic infection. Taking the clinical evaluation as indicating the presence of herpetic infection, the nested polymerase chain reaction and culture had respective sensitivities of 19/26 (73%) and 12/26 (46%) (Chi2 p = 0.02). There was no significant difference in specificities between nPCR62/64 (97%) and culture 63/64 (98%) (Chi2 p = 1.0). Cell content was important for viral detection by nPCR (Chi2 p = 0.07) but not culture. Nesting was found necessary for sensitivity and did not reduce specificity. Assay under-performance appeared related to sub-optimal cell content (20%) but may have reflected clinical over-diagnosis. The results suggest the need for validating specimen cell quality. CONCLUSIONS: This study questions the value of routine laboratory confirmation of mucosal herpetic infection. The adoption of a more discriminatory usage of laboratory diagnostic facilities for genital herpetic infection, taking account of cell content, and restricting it to those cases where it actually affects patient management, may be warranted. (+info)
Characterization of Chlamydia trachomatis omp1 genotypes among sexually transmitted disease patients in Sweden.
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A method for detection and genotyping of genital Chlamydia trachomatis infections based on omp1 gene amplification and sequencing was developed. DNA was extracted from urogenital or urine samples using a Chelex-based method, and an approximately 1,100-bp-long fragment from the omp1 gene was directly amplified and sequenced. Genotyping was performed by BLAST similarity search, and phylogenetic tree analysis was used to illustrate the evolutionary relationships between clinical isolates and reference strains. The method was used to determine the genotypes of C. trachomatis in 237 positive urogenital and/or urine specimens collected at a Swedish sexually transmitted disease clinic during 1 year. The most common genotypes corresponded to serotypes E (47%) and F (17%). The omp1 gene was highly conserved for genotype E (106 of 112 samples without any mutation) and F (41 of 42 samples without any mutation) strains but appear slightly less conserved for genotypes G (n = 6) and H (n = 6), where the sequences displayed one to four nucleotide substitutions relative to the reference sequence. Genotyping of samples collected at the follow-up visit indicated that two patients had become reinfected, while three other patients suffered treatment failure or reinfection. One woman appeared to have a mixed infection with two different C. trachomatis strains. This omp1 genotyping method had a high reproducibility and could be used for epidemiological characterization of sexually transmitted Chlamydia infections. (+info)
Urogenital distribution of a mouse membrane-associated prostaglandin E(2) synthase.
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PGE(2) plays a critical role in regulating renal function and facilitating reproduction. One of the rate-limiting biosynthetic enzymes in PGE(2) synthesis is the terminal PGE(2) synthase (PGES). In the present studies, we report the functional expression of a membrane-associated murine PGES (mPGES) and its expression in urogenital tissues. Two independent cDNA clones sharing an identical open reading frame of 459 bp and encoding a peptide of 153 amino acids, but differing in the 3'-untranslated region, were identified. Assays for enzymatic activity, using microsomes prepared from cells transfected with mPGES cDNA, showed that these cDNA sequences encode a functional protein that catalyzes the conversion of PGH(2) to PGE(2). Constitutive expression of mPGES was highest in the mouse kidney, ovary, and urinary bladder but was also expressed at lower levels in uterus and testis. Renal mPGES expression was predominantly localized to epithelia of distal tubules and medullary collecting ducts. High expression was also seen in transitional epithelial cells of bladder and ureter and in the primary and secondary follicles in the ovary. In conclusion, mPGES is constitutively expressed along the urogenital tract, where it may have important roles in normal physiology and disease. (+info)