Nucleoli, rRNA genes and ITS region in Posidonia oceanica (L.) Delile.

The maximum number of nucleoli was counted in interphase nuclei of Posidonia oceanica, and a restriction pattern of nuclear rDNA was obtained after digestion with four restriction endonucleases and Southern hybridization. P. oceanica has only one type of ribosomal gene whose size was estimated to be 18.5 kbp long. The nucleotide sequence of the entire ITS region was also determined by direct sequencing of PCR amplified DNA fragments. The sequence of the ITS region was aligned with those of homologous regions of other monocots available in literature, and phylogenetic trees were obtained.

Characterization of the aldolase B intronic enhancer.

The aldolase B gene is transcribed at a high level in the liver, kidney, and small intestine. This high level of gene expression results from cooperation between a weak but liver-specific promoter and an intronic activator. A deletional study of this activator present in the first intron allowed us to ascribe the maximal enhancer function to a 400-base pair (bp) fragment (+1916 to + 2329). This enhancer is highly liver-specific and enhances the activity of heterologous minimal promoters in a position and distance-independent fashion in transiently transfected Hep G2 hepatoma cells. The aldolase B enhancer is composed of two domains, a 200-bp module (Ba) inactive by itself but which synergizes with another 200-bp module (Bb) that alone retains 25% of the total enhancer activity. The Bb sequence is 76% homologous between human and rat genes and contains several binding sites for liver-enriched nuclear factors. By electrophoretic mobility shift assays, we demonstrated that elements 5 and 7 bind hepatic nuclear factor 1 (HNF1), whereas element 2 binds hepatic nuclear factor 4 (HNF4). A functional analysis of the enhancer whose elements have been mutated demonstrated that mutation of any of the HNF1 sites totally suppressed enhancer activity, whereas mutation of the HNF4-binding site reduced it by 80%.

Symmetrical periumbilical extension of a midline incision: a simple technique.

BACKGROUND: Periumbilical extension of midline incision often results in an irregular, unaesthetic scar with beveled edges. TECHNIQUE: An Allis clamp is placed at the lateral margin of the umbilicus with subsequent medial traction. This straightens the proposed periumbilical incision, resulting in a symmetrical scar. EXPERIENCE: We have used this technique extensively over the past several years with excellent results and no adverse sequelae. CONCLUSION: This simple technique results in the symmetrically curvilinear, nonbeveled periumbilical extension of a midline incision.

Endocervical curettage, cone margins, and residual adenocarcinoma in situ of the cervix.

OBJECTIVE: To evaluate endocervical curettage (ECC) and cone margin involvement in the management of adenocarcinoma in situ of the cervix. METHODS: Forty-two women with adenocarcinoma in situ without any associated invasive component underwent 49 cervical conizations. The ECC, cone margin involvement, and residual endocervical glandular disease were evaluated in a retrospective descriptive study. RESULTS: The patients ranged from 18 to 65 years old, with a median of 34 years and a mean of 37 years. Nineteen of 42 (45%) of the women presented with initial cervicovaginal cytology suggesting endocervical glandular abnormality. Twenty-seven patients (64%) had mixed lesions of adenocarcinoma in situ and squamous dysplasia noted in their cervical biopsy, conization, or hysterectomy specimens. Forty ECCs were performed at colposcopy or immediately after conization; 28 patients with ECCs subsequently underwent conization, and 12 underwent hysterectomy. Residual adenocarcinoma in situ was found in 18 (67%) of the 27 patients with negative ECCs and in ten of 13 women with positive ECCs. Residual adenocarcinoma in situ was found in two of seven patients with negative cone margins and seven of ten patients with positive margins. CONCLUSION: We found that negative ECCs and uninvolved cone margins in patients with cervical adenocarcinoma in situ were not reassuring of the absence of residual endocervical glandular disease in subsequent surgical specimens. Conservative management and subsequent surveillance of adenocarcinoma in situ should be undertaken with caution.

An intronic enhancer essential for tissue-specific expression of the aldolase B transgenes.

Expression in mice of transgenes directed by regulatory regions of the rat aldolase B gene requires the presence of a B element located in the first intron, while constructs devoid of this intronic enhancer are silent. Histo- and immunochemical staining of transgenic tissue sections showed that the longer transgene was expressed in the proximal tubular cells of the kidney, enterocytes located in small intestine villi and liver parenchymal cells. In the liver, a maximal expression was observed in perivenous hepatocytes, while the transgene was weakly active in periportal hepatocytes, which reproduced the pattern of functional zonation already reported for other glycolytic and gluconeogenic genes in the liver. We also established that the transgene retained the necessary elements for a correct chronological expression during development but was lacking elements necessary for activation by high carbohydrate diet. Instead, transgene expression was paradoxically stimulated in fasted animals, suggesting that the endogenous gene, which must be active under both glycolytic and gluconeogenic conditions, could possess distinct elements activating it in fasted as well as in carbohydrate-fed animals; the former element might be conserved in the transgene and the latter one might be lost.

Bolus mitomycin C and 5-FU with sequential radiation for poor-prognosis locally advanced cervical cancer.

Forty patients with locally advanced cervical carcinoma were entered into a protocol utilizing the bolus administration of both mitomycin C (10 or 15 mg) on Day 1 and 5-fluorouracil (400 mg) on Day 1-5 followed by sequential pelvic irradiation on Day 6 between September 1980 and October 1985. All patients had poor-prognosis FIGO stage IB, IIB, IIIB, or IVA disease. Only patients with poor prognosis factors such as bulky tumor masses of 5 cm or greater noted on the initial physical exam (37 patients) or poorly differentiated histology (3 patients) were eligible for this study. There were three severe side effects seen in the 24 patients receiving 15 mg mitomycin C. One patient developed thrombocytopenia, one patient developed acute radiation enteritis, and the third patient developed radiation proctitis requiring laser therapy. Only 1 of 16 patients receiving 10 mg mitomycin C developed a complication (thrombocytopenia). Neutropenia was mild in all patients. No infections were seen. Thrombocytopenia never warranted platelet transfusion. No patients developed therapy-related bowel obstruction or fistulae. Median follow-up was 11.3 years with a range of 6.2-14.2 years. A complete response rate of 63%, a local control rate of 58%, and a 5-year survival rate of 44% were obtained. This does not appear to offer any benefit over radiation alone. This present study supports the superiority of higher dose concurrent infusional chemotherapy and radiation over low-dose sequential bolus chemotherapy and radiation.

Mature cystic teratoma: a clinicopathologic evaluation of 517 cases and review of the literature.

OBJECTIVE: To evaluate the clinical and pathologic presentation of mature cystic teratomas and the trends in management over a 14-year study period. METHODS: Tumor registry data and medical records between January 1, 1975 and December 31, 1989 were analyzed with respect to patient age, tumor size, bilaterality, malignant transformation, and treatment. RESULTS: Five hundred seventy-three tumors were removed from 517 patients. The median and mean (+/- standard deviation) age was found to be 30 and 32 +/- 11.3 years, respectively. Three hundred ten (60%) of the patients were asymptomatic. The mean tumor size was 6.4 +/- 3.5 cm. The bilaterality rate was 10.8%. The rate of torsion was 3.5%; larger tumors underwent torsion more frequently than smaller tumors (P = .029). The rate of malignant transformation was 0.17%. The mean cyst diameter for patients undergoing cystectomy was 5.7 +/- 2.4 cm; for oophorectomy, 8.0 +/- 4.1 cm; and for hysterectomy, 6.1 +/- 3.8 cm. Oophorectomies were performed for larger tumors when compared to cystectomies (P = .01). The number of hysterectomies was stable throughout the study period, whereas the number of oophorectomies decreased and the number of cystectomies increased markedly. Contralateral ovarian biopsy was common (48.5%) early in the study period. By 1989, the biopsy rate was less than 1%. CONCLUSIONS: We found the prevalence rates of symptomatic tumors, torsion, and malignant degeneration to be less than those previously reported by most other investigators. In addition, there has been an important change over the past 14 years in the management of these neoplasms, with an increased tendency for ovarian preservation, as evidenced by the more frequent use of cystectomy and a decrease in contralateral ovarian biopsy.

Stage II adenocarcinoma of the endometrium treated by two standard regimens of combined preoperative irradiation and surgery.

We retrospectively analyzed 77 patients with stage II endometrial carcinoma treated with standard regimens of preoperative radiotherapy (RT) and surgery (S). The age range was 31-74 years with a median of 56.3 years. Thirty-three patients received 40 Gy whole pelvis RT followed by either radical or modified radical hysterectomy. Forty-four patients received 50 Gy whole pelvis RT and sequential intrauterine and intravaginal cesium-137 brachytherapy followed by a simple hysterectomy. Median follow-up was 111 months. No patient was lost to follow-up. The overall 5-year actuarial survival was 78%. There was no significant difference between the two treatment groups. Several prognostic variables were analyzed. Those with histologic grade I and II had 5-year survival of 89% and 83%, respectively, compared to 62% for grade III (P =0.045). The 5-year survival for microscopic cervical involvement was 87% compared to 59% for gross involvement (P= 0.008). Patients with negative or microscopic residual tumor in the surgical specimen and those with negative lymph nodes had less risk of treatment failure. Local failure occurred in only 9%. Major complications (3%) were seen only in the radical surgery group. Combined preoperative RT and S provide high cure rates with minimal complications for patients with stage II endometrial carcinoma. Patients with adverse prognostic factors are candidates for trials of more aggressive local and systemic therapy.

Activity of the rat liver-specific aldolase B promoter is restrained by HNF3.

Although it contains binding sites for HNF1, NFY and C/EBP/DBP, the proximal promoter of the aldolase B gene is surprisingly weak when tested by transient transfection in differentiated hepatoma cells. This low activity could be due to overlapping between HNF1 and HNF3 binding sites in element PAB, from -127 to -103 bp with respect to the cap site. Replacement of the PAB region by a consensus HNF1 binding site unable to bind HNF3, results in a 30 fold activation of the promoter, in accordance with the hypothesis that activity of the wild-type promoter is normally restrained by HNF3 binding to PAB competitively with HNF1. Consistently, transactivation of the wild-type promoter by excess HNF1 is very high, most likely due to the displacement of HNF3, while the construct with the exclusive HNF1 binding site is weakly transactivated by HNF1. The inhibitory effect of HNF3 on HNF1-dependent transactivation is clearly due to competition between these two factors for binding to mutually exclusive, overlapping sites; indeed, when HNF1 and HNF3 sites are contiguous and not overlapping, the resulting promoter is as active as the one containing an exclusive HNF1 binding site. A construct in which PAB has been replaced by an exclusive HNF3 binding site is weakly expressed and is insensitive to HNF3 hyperexpression. DBP-dependent transactivation, finally, is independent of the nature of the element present in the PAB region.

Wilms' tumor of the uterus. A case report.

Extrarenal Wilms' tumors (nephroblastomas) are considered rare, with only 36 cases reported to date. A primary Wilms' tumor of the uterus has been reported on two previous occasions. A third case is presented and the histologic features and histogenesis of the tumor discussed.

Competition between transcription factors HNF1 and HNF3, and alternative cell-specific activation by DBP and C/EBP contribute to the regulation of the liver-specific aldolase B promoter.

The aldolase B proximal promoter is controlled by at least five elements spanning from -190 to -103 bp with respect to the start site of transcription. From 5' to 3', we found: a negative DE element, an activating C/EBP-DBP binding site, a CCAAT box binding NFY that seems to play a negative role, and an activating element consisting of two overlapping binding sites for HNF-1 and HNF-3. Contransfection experiments of aldolase B/CAT constructs and of expression vectors for different transcription factors were carried out in human hepatoma Hep G2 cells. We found that DBP and HNF-1 are strong transactivators of the aldolase B promoter while C/EBP and vHNF-1 are only weak activators and HNF-3 alone does not modify such activity. Deletion of the distal negative element results in a similar transactivation by C/EBP and DBP, enhanced for the former and reduced for the latter. In hepatocytes in primary culture, the strong transactivator is C/EBP while DBP is essentially inactive. This tissue-specificity of C/EBP and DBP action could depend on interaction with tissue-specific proteins bound to a neighbouring site, probably DE. Finally, HNF3 behaves as a very strong anti-activator of the aldolase B promoter. It competitively antagonizes transactivation by HNF-1 and non-competitively transactivation by DBP. This negative effect of HNF-3 and tissue-specificity of the transactivation potential of DBP and C/EBP are unique features of the aldolase B promoter.

Interplay of an original combination of factors: C/EBP, NFY, HNF3, and HNF1 in the rat aldolase B gene promoter.

The rat aldolase B 5' flanking region (nucleotides - 194 to +41) contains sufficient information for liver-specific expression. A detailed investigation of factors binding to the rat aldolase B 5' flanking region has allowed us to identify three distinct factors that filled different sites of this region (A, B, C). The liver-enriched C/EBP or related factors bind to box C, as demonstrated by the specific interaction with bacterially expressed C/EBP protein. Box B bearing the CCAAT sequence binds the ubiquitous factor NFY. Surprisingly, Box A is able to bind two liver enriched factors, namely HNF1 and HNF3. However, in the context of the intact promoter, as shown by footprinting competition experiments, HNF3 binds solely to this sequence. HNF3, but not HNF1 is a transcriptional activator as demonstrated in the in vitro transcription assay.

Expression of the rat aldolase B gene: a liver-specific proximal promoter and an intronic activator.

The nature and location of the cis-acting DNA sequences regulating expression of the rat aldolase B gene has been investigated. Two liver-specific DNAse I hypersensitive sites were detected, one located just upstream from the cap site, the second in the middle of the first, 4.8-kbp-long, intron. A fragment of 190 bp 5' to the cap site behaved as a tissue-specific but weak core promoter: it directed a detectable reporter gene expression in the Hep G2 cells and hepatocytes, but not in fibroblasts. The tissue-specific expression was stimulated at least 16 fold when constructs contained the entire first intron. The intronic activating sequences could be ascribed to an inner 2 kbp fragment in which the downstream liver-specific DNAse I hypersensitive site was located.

[Predominant position of impacted lower third molars]. / Aferição das posições prevalentes dos terceiros molares inferiores inclusos.

This research tries to establish the predominant position of the included mandibular third molar giving emphasis to the surgical planning. The results were in agreement to that in world literature.

Molecular analysis of aldolase B genes in hereditary fructose intolerance.

The molecular basis of hereditary fructose intolerance (HFI) was studied in 50 subjects (41 pedigrees, 82 apparently independent mutant alleles of aldolase B) by direct analysis of aldolase B genes amplified by means of the polymerase chain reaction. The mutation A149P (ala 149----pro) was found in 67% of alleles but was significantly more common in patients from northern than from southern Europe. Two other point mutations of aldolase B were identified. A174D (C----A; ala 174----asp) was found in subjects from Italy, Switzerland, and Yugoslavia (overall frequency 16%) but not in those from the United Kingdom, France, or the United States. L288 delta C carried a single base-pair deletion causing frameshift at codon 288 and was restricted to Sicilian subjects. By testing for these mutations in amplified DNA with a limited panel of allele-specific oligonucleotides, more than 95% of HFI patients will be susceptible to genetic diagnosis.

Mitral valve prolapse and thromboembolic disease in pregnancy: a case report.

Mitral valve prolapse is usually a benign condition, however, serious complications have been reported to be associated with it. A report of retinal artery occlusion associated with mitral valve prolapse and pregnancy is presented.

Characterization of three optional promoters in the 5' region of the human aldolase A gene.

We undertook cloning and sequencing of the 5' portion of the human aldolase A gene to elucidate the mechanisms that govern synthesis of its different mRNAs. The sequenced gene is the only active gene in human-rodent fibroblastic somatic hybrids, while the other aldolase A-related sequences are inactive. S1 mapping and primer extension analysis enabled us to demonstrate that three promoter regions were implicated in the initiation of different aldolase A mRNAs, differing only in their 5' non-coding extremities. A distal promoter, N (non-specific), governs the synthesis of a 5' non-coding region of 142 bases composed of two exons, N1 and N2, which are found in a variety of tissues. A median promoter, M (muscle), is only active in skeletal muscle, and initiates the transcription by a 5' non-coding exon of 45 bases. Finally, a proximal promoter, H (housekeeping), contained in a "G + C-rich island", permits transcription of three colinear mRNAs containing 172, 126 or 112 bases of 5' non-coding sequence; their expression seems ubiquitous. These three promoters are arranged in 1.5 X 10(3) base-pairs of DNA. Homologies between rat and human genomic sequences and the absence of homology between promoters or 5' non-coding exons of the same species exclude a recent duplication of the promoter regions.